US20040001801A1 - Conjugates activated by cell surface proteases and therapeutic uses thereof - Google Patents
Conjugates activated by cell surface proteases and therapeutic uses thereof Download PDFInfo
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- US20040001801A1 US20040001801A1 US10/156,214 US15621402A US2004001801A1 US 20040001801 A1 US20040001801 A1 US 20040001801A1 US 15621402 A US15621402 A US 15621402A US 2004001801 A1 US2004001801 A1 US 2004001801A1
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- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- A61K38/19—Cytokines; Lymphokines; Interferons
- A61K38/193—Colony stimulating factors [CSF]
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- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- A61K38/19—Cytokines; Lymphokines; Interferons
- A61K38/20—Interleukins [IL]
- A61K38/2006—IL-1
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- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- A61K38/19—Cytokines; Lymphokines; Interferons
- A61K38/20—Interleukins [IL]
- A61K38/2013—IL-2
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- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- A61K38/19—Cytokines; Lymphokines; Interferons
- A61K38/20—Interleukins [IL]
- A61K38/204—IL-6
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/43—Enzymes; Proenzymes; Derivatives thereof
- A61K38/46—Hydrolases (3)
- A61K38/48—Hydrolases (3) acting on peptide bonds (3.4)
- A61K38/49—Urokinase; Tissue plasminogen activator
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- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
- C12N9/50—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25)
- C12N9/64—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from animal tissue
- C12N9/6421—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from animal tissue from mammals
- C12N9/6424—Serine endopeptidases (3.4.21)
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Definitions
- Conjugates, compositions and methods for localized delivery of therapeutic agents for treating a variety of disorders such as, proliferative diseases, autoimmune diseases, infectious diseases and inflammatory diseases.
- the conjugates which act as prodrugs, contain therapeutic agents and peptidic substrates that are cleaved by cell surface proteases to release therapeutic agents in the vicinity of the targeted cells.
- chemotherapeutic agents typically systemic administration.
- chemotherapeutic agents are cytotoxic agents that act by inhibiting proliferation or other metabolic processes, so that actively proliferating and growing cells will be targeted by the agent.
- targeting is not highly specific, and the side-effects are often devastating.
- conjugates that contain a targeting agent, such as an antibody and/or growth factor, and a therapeutic agent, that act on specific cells; the use of antisense technology that is targeted to specific genes and/or proteins; the use of genetic therapy to provide, for example, correct copies of defective genes or pharmaceutically active compounds, and the use of toxins that are relatively non-toxic unless delivered intracellularly.
- a targeting agent such as an antibody and/or growth factor
- a therapeutic agent that act on specific cells
- antisense technology that is targeted to specific genes and/or proteins
- genetic therapy to provide, for example, correct copies of defective genes or pharmaceutically active compounds
- toxins that are relatively non-toxic unless delivered intracellularly.
- the compounds are conjugates that contain a peptidic substrate for a cell surface protease, or a soluble, shed or released form thereof, and an agent that upon cleavage by the protease is a therapeutic agent or in a form that can be activated by the targeted cell or tissue or in the localter thereof.
- the agents include therapeutic agents, such as a cytotoxic agents, drugs, therapeutic nucleic acid moleulces, and diagnostic agents, such as labelled moieties and imaging agents.
- the conjugates contain one or more substrates for one or a plurality of cell surface proteases linked either directly or via a linker to a targeted agent, including a therapeutic agent, such as a cytotoxic agent.
- the conjugates provided herein contain the following components: (peptidic substrate) s , (linker) q , and (targeted agent) t in which: at least one peptidic substrate moiety is linked with or without a linker (L) to at least one therapeutic agent, s is 1 or more and each substrate is the same or different, and is typically is between 1 and 6, generally 1, 2 or 3; q is 0 or more as long as cell surface protease(s) cleaves the peptidic substrate(s) and releases active therapeutic agent or, releases the agent in a form that is converted by the cell, tissue or surrounding environment to an active form, q is 0 to t, generally 1 to 4; t is 1 or more, generally 1 or 2 and each targeted agent are the same or different; linker refers to any link
- the therapeutic agents include any biologically active molecule. These agents include toxins, cytokines and lymphokines, growth factors, nucleic acid molecules, such as antisense nucleic acid, dsRNA, and DNA molecules.
- the therapeutic agents include those that are active intracellularly, such as cytotoxins, or extracellularly, such as modulators of the activity of extracellular receptors. When in the conjugates the therapeutic agents are substantially inactive, and when cleaved are released in active form or in a form that can be activated by the targeted cell or tissue or environment thereof.
- peptide l is a peptidic substrate for a cell surface protease; s is greater than or equal to 1, or is 1 to 6, or is 1 or 2, or is 1; linker is any linker; q is greater than or equal to 0, or is 0 to 4, or is 0 or 1; the therapeutic agent is, for example, a cytotoxic agent, including, but not limited to, an anti-tumor, anti-angiogenic, anti-cancer, pro-apoptotic and anti-mitotic agents; and t is 1 or more, or is 1 or 2.
- the therapeutic agent is covalently attached, optionally via a linker L, to either the C-terminus or the N-terminus of the peptidic substrate.
- peptide l is a substrate for a cell surface protease whereby, upon action of the protease, the conjugate, which is substantially inactive, is cleaved at a point on the peptidic substrate chain to release a compound of the formula:
- the therapeutic agent is, for example, a cytotoxic agent
- peptide a is a truncated version of peptide l resulting from cleavage at the P1-P1′ bond.
- the conjugates can be used to target and deliver the targeted agents to specific cells, and hence can be used for the treatment any diseases that are associated with cells or tissues that express a cell surface protease, including cell-associated and cell-localized proteases.
- the cells on which or near which such proteases are expressed are not necessarily involved in the disease or disease process, but are present and can serve to present the protease, which cleaves the targeted conjugate.
- Methods of treatment of diseases associated with cells or tissues that express a cell surface protease including cell-associated and cell-localized proteases.
- the diseases include, but not limited to, proliferative diseases, autoimmune diseases, infectious diseases and inflammatory diseases.
- diseases include e, but are not limited to, rheumatoid arthritis, lupus, multiple sclerosis, psoriasis, diabetic retinopathies, other ocular disorders, including recurrence of pterygii, scarring excimer laser surgery and glaucoma filtering surgery, various disorders of the anterior eye, cardiovascular disorders, restenosis, chronic inflammatory diseases, wounds, circulatory disorders, crest syndromes, bacterial infections, viral diseases, including AIDS, dermatological disorders, and cancer, including solid neoplasms and vascular tumors, including, but are not limited to, lung, colon, esophageal, breast, ovarian and prostate cancers.
- the methods involve identifying cell-surface protease-associated disease by identifying a cell involved in the disease process or a cell in the vicinity of the cell involved in the disease process; and identifying a cell surface protease on the cell. Conjugates that target such proteases as provided herein can then be prepared.
- FIG. 1 shows exemplary doxorubicin conjugates provided herein and in vitro CT 50 (min) data for cleavage of the conjugates by MTSP1.
- FIG. 2 shows exemplary doxorubicin conjugates provided herein and in vitro CT 50 (min) data for cleavage of the conjugates by u-PA.
- FIG. 3 shows exemplary taxol conjugates provided herein and in vitro CT 50 (min) data for cleavage of the conjugates by MTSP1.
- FIG. 4 shows exemplary taxol conjugates provided herein and in vitro CT 50 (min) data for cleavage of the conjugates by u-PA.
- FIG. 5 shows exemplary doxorubicin and taxol conjugates provided herein and in vitro CT 50 (min) data for cleavage of the conjugates by ET1 (endotheliase 1).
- a targeted agent is any agent intended for targeted delivery and includes therapeutic agents and diagnostic agents and any other agent intended for targeted delivery.
- targeted delivery means delivery to a selected cell or tissue that expresses a protease that releases the targeted agent. Such delivery does not have to be exclusively to such selected cell or tissue, but must include it, and generally deliveries higher amounts to such selected cells or tissues. Delivery includes introduction into a cell or tissue or binding to the cell or tissue or release in the vicinity of the cell or tissue. For example, in some instances, a tumor induces production of proteases, receptors, co-factors or substrates asssociated with the stroma; delivery, thus, includes targeting such induced stromal activities, such as proteases, receptors and/or enzyme co-factors, in invading cells or cells in the tumor that is targeted.
- therapeutic index is the ratio of LD 50 /ED 50 .
- a therapeutic agent is any drug or other agent that is intended for delivery to a targeted cell or tissue, such as proliferating cells, including tumor cells and cells involved in a proliferative, typically an undesirable, response.
- Therapeutic agents include, but are not limited to, anti-cancer agents, anti-angiogenic agents, pro-apoptotic agents, anti-mitotic growth factors, cytokines, such as tumor necrosis factors and interleukins, and cytotoxic agents and other such agents as described herein and known to those of skill in the art.
- Therapeutic agents include those that are active upon internalization and also those that act extracellularly, such modulators of the activities of certain cell surface receptors, such as G proteins that transduce extracellular signals.
- an inactive therapeutic agent is a therapeutic agent that is conjugated to a peptide and thereby, either by virtue of conformational changes or size or other factors such as steric hinderance does not exhibit any or exhibits substantially reduced activity compared to the released active therapeutic agent.
- conjugated doxorubicin is not toxic to cells until it is released from the conjugate in a form that can enter the cell. Upon cleavage of the agent from the conjugate it is in active form or in a form that is further processed by one or a plurality of steps, including enzymatically or chemically, in or on the cell, into an active form.
- an active therapeutic agent is a therapeutic agent that has been released from the conjugate by cleavage of the peptidic substrate portion of the conjugate.
- the active therapeutic agent is by virtue of cleavage able to exhibit its intended activity, typically by entering the cell.
- the therapeutic agents When conjugated the therapeutic agents have reduced or no activity as therapeutic agents, and upon cleavage are released in the vicinity of a cell.
- an anti-cancer agent refers to any agents used in the treatment of cancer. These include any agents, when used alone or in combination with other compounds, that can alleviate, reduce, ameliorate, prevent, or place or maintain in a state of remission of clinical symptoms or diagnostic markers associated with neoplasm, tumor or cancer, and can be used in methods, combinations and compositions provided herein.
- Non-limiting examples of anti-neoplasm agents include anti-angiogenic agents, alkylating agents, antimetabolite, certain natural products, platinum coordination complexes, anthracenediones, substituted ureas, methylhydrazine derivatives, adrenocortical suppressants, certain hormones, antagonists and anti-cancer polysaccharides.
- substantially inactive with reference to the conjugated thereapeutic agent means at least 1%, generally 10, 20, 30, 50, 60, 70, 80 or 90 or 100% inactive compared to the unconjugated therapeutic agent in a standard or art-recognized assays, such as in vitro or in vivo assays, that assess the therapeutic activity of the agent.
- a targeted cell or tissue refers to the cells or tissues that include cell surface proteases that cleave the conjugates.
- the cells or tissues can be involved in the disease or can be present at the disease loci or locus by virtue of participation in the disease process or merely serendipitously.
- angiogenesis is intended to broadly encompass the totality of processes directly or indirectly involved in the establishment and maintenance of new vasculature (neovascularization), including, but not limited to, neovascularization associated with tumors.
- anti-angiogenic treatment or agent refers to any therapeutic regimen and compound, that, when used alone or in combination with other treatment or compounds, can alleviate, reduce, ameliorate, prevent, or place or maintain in a state of remission, one or more clinical symptoms or diagnostic markers associated with undesired and/or uncontrolled angiogenesis.
- an anti-angiogenic agent refers to an agent that inhibits the establishment or maintenance of vasculature.
- agents include, but are not limited to, anti-tumor agents, and agents for treatments of other disorders associated with undesirable angiogenesis, such as diabetic retinopathies, hyperproliferative disorders and others.
- non-anti-angiogenic anti-tumor agents refer to anti-tumor agents that do not act primarily by inhibiting angiogenesis. Whether anti-tumor agents act primarily by inhibiting angiogenesis can be determined using the assays provided herein, or using other assays well known to those of skill in the art.
- undesired and/or uncontrolled angiogenesis refers to pathological angiogenesis wherein the influence of angiogenesis stimulators outweighs the influence of angiogenesis inhibitors.
- deficient angiogenesis refers to pathological angiogenesis associated with disorders where there is a defect in normal angiogenesis resulting in aberrant angiogenesis or an absence or substantial reduction in angiogenesis.
- a cell surface protease is any protease that is located on or at a cell surface and/or proteases that are located at the cell surface by virtue of a specific binding interaction with a receptor therefor, or that is localized at or near or associated with the cell surface.
- An exemplary protease located at the cell surface by virtue of a specific binding interaction with a receptor therefor is urokinase plasminogen activator (u-PA) bound to urokinase plasminogen activator receptor (u-PAR).
- u-PA urokinase plasminogen activator
- u-PAR urokinase plasminogen activator receptor
- cell surface proteases contemplated herein include cell surface-associated proteases. It also includes all forms thereof that can be circulating or inside a cell.
- Cell surface protease To be categorized as a cell surface protease, there must be at least one form thereof that is located (i.e. on the surfaces such as transmembrane protease or bound to receptor therefor) on the surface of a cell at some point in its cycle.
- Cell surface protease include serine proteases, such as, but are not limited to, the transmembrane serine protease (MTSPs) and endotheliases and urokinases.
- MTSPs transmembrane serine protease
- a serine protease refers to a diverse family of proteases in which a serine residue is involved in the hydrolysis of proteins or peptides.
- the serine residue can be part of the catalytic triad mechanism, which includes a serine, a histidine and an aspartic acid in the catalysis, or be part of the hydroxyl/ ⁇ -amine or hydroxyl/ ⁇ -amine catalytic dyad mechanism, which involves a serine and a lysine in the catalysis.
- SPs of mammalian, including human, origin are particularly interested in these SPs of mammalian, including human, origin.
- soluble and released forms of cell surface proteases are contemplated. Such forms include, for example, forms found in serum upon proteolytic degradation or other removal of the extracellular portion of membrane bound protease, and splice variants that do not include a transmembrane domain.
- the protease activity of cell surface proteases and proteases associated with cells can be exploited to provide a means to concentrate therapeutic agents, such as cytotoxic agents, near such cells by providing conjugates that are activated upon cleavage by such enzymes.
- Such conjugates upon the action of a cell surface protease or cell-associate protease, release the therapeutic agent, such as a cytotoxic agent, or a derivative thereof that can be converted to a therapeutic agent, locally at the site of action.
- transmembrane serine protease refers to a family of transmembrane serine proteases that share common structural features as described herein (see, also Hooper et al. (2001) J. Biol. Chem .276:857-860).
- MTSP encompasses all proteins encoded by the MTSP genes, including but are not limited to: MTSP1, MTSP3, MTSP4, MTSP6, MTSP7, MTSP9, MTSP10, MTSP12, MTSP20, MTSP22 and MTSP25 or an equivalent molecule obtained from any other source or that has been prepared synthetically or that exhibits the same activity.
- MTSPs include, but are not limited to, corin, enteropeptidase, human airway trypsin-like protease (HAT), TMPRSS2 and TMPRSS4.
- HAT human airway trypsin-like protease
- TMPRSS2 TMPRSS4.
- the MTSPs described herein can be used to identify other MTSPs.
- Methods for isolating nucleic acid encoding other MTSPs including nucleic acid molecules encoding full-length molecules and splice variants and MTSPs from species, such as cows, sheep, goats, pigs, horses, primates, including chimpanzees and gorillas, rodents, dogs, cats and other species of interest, such as domesticated animals, farm and zoo animals are known to those of skill in the art and are outlined herein.
- nucleic acid molecules described herein including those set forth in SEQ IDs can be used to obtain nucleic acid molecules encoding full-length MTSP polypeptides from human sources or from other species, such as by screening appropriate libraries using the nucleic acid molecules or selected primers or probes based thereon.
- Sequences of encoding nucleic acid molecules and the encoded amino acid sequences of exemplary MTSPs and/or domains thereof are set forth in SEQ ID Nos. 1-45, 269-270 and 272-276.
- the term also encompasses MTSPs with amino acid substitutions that do not substantially alter activity of each member and also encompasses polyeptides encoded by splice variants thereof.
- MTSPs with amino acid substitutions such that the resulting polypeptide retains at least 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% of the proteolytic activity of the unaltered polypeptide, and also encompasses MTSPs encoded by splice variants thereof and MTSPs encoded by allelic variants, such as single nucleotide polymorphisms (SNPs).
- SNPs single nucleotide polymorphisms
- Suitable substitutions including, although not necessarily, conservative substitutions of amino acids, are known to those of skill in this art and can be made without eliminating the biological activity, such as the catalytic activity, of the resulting molecule.
- MTSPs include those of animal, such as mammalian, including human, origin.
- a “protease domain of an MTSP” refers to an extracellular protease domain of an MTSP that exhibits proteolytic activity and shares homology and structural features with the chymotryp-sin/trypsin family protease domains. Hence it is at least the minimal portion of the domain that exhibits proteolytic activity as assessed by standard in vitro assays. Contemplated herein are such protease domains and catalytically active portions thereof.
- Exemplary MTSP polypeptides are set forth in SEQ ID Nos. 1-45, 269-270 and 272-276, and including smaller portions thereof that retain or exhibit protease activity.
- the protease domains vary in size and constitution, including insertions and deletions in surface loops. They retain conserved structure, including at least one of the active site triad, primary specificity pocket, oxyanion hole and/or other features of serine protease domains of proteases.
- the protease domain is a portion of a MTSP, as defined herein, and is homologous to a domain of other MTSPs.
- MTSPs include, MTSP1, MTSP3, MTSP4, MTSP6, MTSP7, MTSP9, MTSP10, MTSP12, MTSP20, MTSP22 and MTSP25 (see SEQ ID Nos. 1-19, 42-45, 269-270 and 272-276; see, also International PCT application No. WO 02/00860 (see SEQ ID Nos. 38 and 97 therein, which provide an MTSP12 variant); corin (SEQ ID Nos. 28 and 29), enteropeptidase (SEQ ID Nos. 30 and 31) human airway trypsin-like protease (HAT) (SEQ ID Nos. 32 and 33), hepsin (SEQ ID Nos.
- TMPRSS2 SEQ ID Nos. 36 and 37
- TMPRSS4 SEQ ID Nos. 38 and 39.
- S1 fold see, e.g., Internet accessible MEROPS data base
- the MTSPs protease domains share a high degree of amino acid sequence identity.
- His, Asp and Ser residues necessary for activity are present in conserved motifs.
- the activation site which results in the N-terminus of second chain in the two chain forms has a conserved motif and readily can be identified (see, e.g., amino acids 801-806, SEQ ID No.
- amino acids 406-410 SEQ ID No. 31; amino acids 186-190, SEQ ID No. 33; amino acids 161-166, SEQ ID No. 35; amino acids 255-259, SEQ ID No. 37; amino acids 190-194, SEQ ID No. 39 and other as known to those of skill and the art and/or as described herein).
- the C 573 (SEQ ID NO. 45 is a free Cys in a single chain form of the protease domain.
- the protease also can be provided as a two chain molecule. Single chain and two chain forms are proteolytically active. A two chain form is produced by bonding, typically between the C 573 and a Cys outside the protease domain, such as Cys 296 . Upon activation cleavage the disulfide bond remains resulting in a two chain polypeptide.
- the size of chain “A” is a function the starting length of the polypeptide prior to activation cleavage between the R 462 and I 463 .
- Two chain forms include at least the protease domain a polypeptide from C 296 up to and including C 573 .
- a two-chain form of the protease domain refers to a two-chain form that is formed from a single chain form of the protease in which the Cys pairing between, e.g., a Cys outside the protease domain such as, for example Cys 573 (SEQ ID No. 45 for MTSP), which links the protease domain to the remainder of the polypeptide, the “A” chain.
- a two chain protease domain form refers to any form in which the “remainder of the polypeptide”, i.e., “A” chain, is shortened and includes a Cys from outside the protease domain.
- the catalytically active domain of an MTSP refers to the protease domain.
- Reference to the protease domain of an MTSP generally refers to a single chain form of the protein. If the two-chain form or both forms is intended, it is so-specified.
- the zymogen form of each protein is a single chain, which is converted to the active two or multi chain form by activation cleavage.
- active form is meant a form active in vivo or in vitro.
- activation cleavage refers to the cleavage of the protease at the N-terminus of the protease domain (generally between an R and I or V in the full-length protein.
- Cys-Cys pairing between a Cys outside the protease domain and a Cys in the protease domain (see, e.g., Cys 573 SEQ ID No. 45
- the resulting polypeptide has two chains (“A” chain and the “B” chain, which is the protease domain of an MTSP).
- Cleavage can be effected by another protease or autocatalytically.
- the conjugates provided herein advantageously contain sites that are recognized by the active cell surface protease (or cell-associated protease) and are cleaved thereby to release active or an inactive prodrug form of a therapeutic agent.
- an MTSP1 whenever referenced herein, includes at least one or all of or any combination of:
- polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 2 or 41;
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 2 or 41; and/or
- the MTSP1 can be from any animal, particularly a mammal, and includes but is not limited to, humans, rodents, fowl, ruminants and other animals.
- the full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- MTSP1 also is referred to TADG-15 and matriptase. As described below, the protein originally designated matriptase appears to be an MTSP1 splice variant or processed product.
- an MTSP3 whenever referenced herein, includes at least one or all of or any combination of:
- polypeptide that comprises the sequence of amino acids set forth as amino acids 205-437 of SEQ ID No. 4;
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 4; and/or
- the MTSP3 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals.
- the full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- an MTSP4 includes at least one or all of or any combination of:
- polypeptide that comprises the sequence of amino acids set forth in any of SEQ ID Nos. 6, 8 or 10;
- polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 6, 8 or 10; and/or
- the MTSP4 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals.
- the full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- an MTSP6 whenever referenced herein, includes at least one or all of or any combination of:
- polypeptide that comprises the sequence of amino acids set forth in any of SEQ ID No. 12;
- polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 12; and/or
- the MTSP6 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals.
- the full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- an MTSP7 includes at least one or all of or any combination of:
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 14; and/or
- the MTSP7 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals.
- the full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- an MTSP9 whenever referenced herein, includes at least one or all of or any combination of:
- polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 18 or 43;
- polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 18 or 270; and/or
- the MTSP9 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals.
- the full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- an MTSP10 whenever referenced herein, includes at least one or all of or any combination of:
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 45; and/or
- the MTSP10 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals.
- the full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- MTSP10 polypeptides including, but not limited to splice variants thereof, and nucleic acids encoding MTSPs, and domains, derivatives and analogs thereof are provided herein.
- Single chain protease domains that have an N-terminus functionally equivalent to that generated by activation of the zymogen form of MTSP10 are also provided.
- the cleavage site for the protease domain of MTSP10 is between amino acid R and amino acids I (R ⁇ IIGGT) (residues 462-467 SEQ ID No. 45).
- an MTSP12 whenever referenced herein, includes at least one or all of or any combination of: SEQ ID No. 19 and 20
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in is set forth as SEQ ID No. 19;
- polypeptide that includes the sequence of amino acids set forth in SEQ ID No. 20 or a catalytically active portion thereof;
- polypeptide that includes a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 20; and/or
- a polypeptide encoded by a splice variant of the MTSP12 that includes the sequence of amino acids set forth in SEQ ID No. 20.
- the MTSP12 polypeptide with the protease domains as indicated in SEQ ID Nos. 19 and 20, is provided.
- the polypeptide is a single or multi-chain polypeptide.
- a protease domain of an MTSP12 whenever referenced herein, includes at least one or all of or any combination of or a catalytically active portion of:
- polypeptide that includes the sequence of amino acids set forth in SEQ ID No. 20 or a catalytically active portion thereof but that does not include the sequence of amino acids set forth in SEQ ID No. 271;
- polypeptide that includes the sequence of amino acids set forth in SEQ ID No. 272 or a catalytically active fragment thereof;
- a polypeptide that includes a sequence of amino acids having at least about 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 20;
- a polypeptide that includes a sequence of amino acids having at least about 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids of the polypeptides of a)-e);
- the MTSP12 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals.
- the full-length zymogen or two-chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two-chain activated form, or a single chain form.
- MTSP12 also includes the variant described International PCT application No. WO 02/00860 (see SEQ ID Nos. 38 and 97 therein).
- an MTSP20 whenever referenced herein, includes at least one or all of or any combination of:
- a polypetide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 274; and/or a polypeptide encoded by a splice variant of the MTSP20 encoded by the sequence of nucleotides that includes the sequence set forth in SEQ ID No. 273.
- the MTSP20 may be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals.
- the full length zymogen or two-chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two-chain activated form, or a single chain form.
- an MTSP22 whenever referenced herein, includes at least one or all of or any combination of:
- a polypetide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 276; and/or
- the MTSP22 may be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals.
- the full length zymogen or two-chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two-chain activated form, or a single chain form.
- an MTSP25 whenever referenced herein, includes at least one or all of or any combination of:
- a polypetide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 270; and/or
- the MTSP25 may be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals.
- the full length zymogen or two-chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two-chain activated form, or a single chain form.
- a human protein is one encoded by nucleic acid present in the genome of a human, including all allelic variants and conservative variations as long as they are not variants found in other mammals.
- PSA prostate specific antigen
- PSA prostate specific antigen
- the relative activity is greater than at least 2:1, 3:1, 4:1, 5:1, 10:1, 50:1 or 100:1.
- activity refers to the ratio k cat /K m , where k cat is the rate of catalytic turnover for a particular enzyme, and K m is the Michaelis constant for the binding of the substrate.
- nucleic acid encoding a protease domain or catalytically active portion of a MTSP shall be construed as referring to a nucleic acid encoding only the recited single chain protease domain or active portion thereof, and not the other contiguous portions of the MTSP as a continuous sequence.
- a CUB domain is a motif that mediates protein-protein interactions in complement components C1r/C1s and has also been identified in various proteins involved in developmental processes.
- a zymogen is an enzymatically inactive protein (i.e, typically, but not necessarily, less than 1% of active form) that is converted to a proteolytic enzyme by the action of an activator, including by autoactivation.
- Inactive means less active than the form those of skill in the art consider to be the active form of the enzyme.
- the ratio of activity of a zymogen to the activated form varies from enzyme-to-enzyme.
- disease or disorder refers to a pathological condition in an organism resulting from, e.g., infection or genetic defect, and characterized by identifiable symptoms.
- the diseases contemplated for treatment herein are any for which a cell surface protease, including a cell-localized or cell-associated protease is asssociated with a targeted cell or tissue involved in the disease or disease process. Such association can be because the protease is involved in the disease or is serendipitously associated with cells involved with the disease. These diseases herein are called cell surface protease-associated diseases.
- a cellsurface protease is identified that is expressed on cells associated with the disorder, such as, for example, immune cells for treating inflammatory diseases, and virally infected cells for treating viral diseases.
- the conjugate is designed as described herein for cleavage by the selected protease.
- neoplasm refers to abnormal new growth, and thus means the same as tumor, which can be benign or malignant. Unlike hyperplasia, neoplastic proliferation persists even in the absence of the original stimulus.
- neoplastic disease refers to any disorder involving cancer, including tumor development, growth, metastasis and progression.
- cancer refers to a general term for diseases caused by any type of malignant tumor.
- malignant as applied to tumors, refers to primary tumors that have the capacity of metastasis with loss of growth control and positional control.
- endotheliase refers to a mammalian protein, including human protein, that has a transmembrane domain and is expressed or active on the surface of endothelial cells and includes a protease domain, particularly an extracellular protease domain, and is generally a serine protease (see, also U.S. application Ser. No. 09/717,473 and International PCT application No. WO 01/36604).
- protease domain particularly an extracellular protease domain
- is generally a serine protease see, also U.S. application Ser. No. 09/717,473 and International PCT application No. WO 01/36604.
- endotheliase encompasses all proteins encoded by the endotheliase gene family, or an equivalent molecule obtained from any other source or that has been prepared synthetically or that exhibits the same activity.
- the endotheliase gene family are transmembrane proteases expressed or active in endothelial cells. These proteases include serine proteases. These include proteins that have these features and also include a protease domain that exhibits sequence homology to the endotheliases 1 and 2. Endotheliase 1 and 2, for example exhibit about 40% or 45% identity. Sequence homology means sequence identity along its length when aligned to maximize identity of at least about 25%, 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% or greater number of residues.
- Sequence homology also is assessed by determining whether the encoding sequences of nucleic acids hybridize under conditions of at least moderate, or for more closely related proteins, high stringency to the nucleic acid molecules provided herein or to those that encode the same proteins but differ in sequence by virtue of the degeneracy of the genetic code.
- endotheliases encompasses endotheliases with amino acid substitutions, including those set forth in Table 1, such that the resulting polypeptide retains at least 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% of the proteolytic activity of the unaltered polypeptide. Suitable substitutions of amino acids are known to those of skill in this art and can be made generally without altering the biological activity of the resulting molecule.
- an endotheliase 1 includes at least one or all of or any combination of:
- polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 22;
- polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 22; and/or
- the endotheliase 1 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals.
- the full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- an endotheliase 2 whenever referenced herein, includes at least one or all of or any combination of:
- polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 24 or 26;
- polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 24 or 26; and/or
- the endotheliase 2 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals.
- the full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- the protease domain of an endotheliase refers to the polypeptide portion of the endotheliase that is the extracellular portion that exhibits protease activity.
- the protease domain is a polypeptide that includes at least the minimum number of amino acids, generally more than 50 or 100, required for protease activity.
- Protease activity can be assessed empirically, such as by testing the polypeptide for its ability to act as a protease. Assays, such as those described in the EXAMPLES, with the exception that a known endotheliase substrate is employed in place of the test compounds, can be used to assess protease activity.
- proteases particularly serine proteases, have characteristic structures and sequences or motifs, the protease domain can be readily identified by such structure and sequence or motif.
- a portion of protease domain of endotheliase refers to a portion of protease domain of endotheliase that is located within or is the extracellular domain of an endotheliase and exhibits serine proteolytic activity. Hence, it is at least the minimal portion of the extracellular domain that exhibits proteolytic activity as assessed by standard assays.
- An exemplary protease domain of an endotheliase is set forth in SEQ ID No. 22 and as amino acids 321-688 and 321-562 of SEQ ID Nos. 24 and 26, respectively. Smaller portions thereof that retain protease activity are contemplated.
- protease domains vary in size and constitution, including insertions and deletions in surface loops. Such domains exhibit conserved structure, including at least one structural feature, such as the active site triad, primary specificity pocket, oxyanion hole and/or other features of serine protease domains of proteases.
- the protease domain is a portion of an endotheliase, as defined herein, but is homologous in terms of structural features and retention of sequence of similarity or homology the protease domain of chymotrypsin or trypsin.
- homologous means about greater than about 25%, 40%, 60%, 80%, 90%, 95%, 98% or greater sequence identity.
- sequence identity the number of conserved amino acids as determined by standard alignment algorithms programs, and used with default gap penalties established by each supplier. Also homology can be assessed by conserved nucleic acid sequence, which includes anything that hybridizes under at least low stringency conditions and encodes the domain.
- nucleic acid sequence alignment programs are commercially available (DNAStar “MegAlign” program (Madison, Wis.) and the University of Wisconsin Genetics Computer Group (UWG) “Gap” program (Madison, Wis.)).
- a polypeptide consists essentially of the protease domain means that the only endotheliase portion of the polypeptide is a protease domain or a catalytically active portion thereof.
- the polypeptide can optionally include additional non-endotheliase-derived sequences of amino acids.
- domain refers to a portion of a molecule, e.g., proteins or nucleic acids, that is structurally and/or functionally distinct from other portions of the molecule.
- an active form of a protease refers to an enzyme that catalyzes hydrolysis of proteins or peptides.
- Reference to a protease includes the active and zymogen or other less active form.
- nucleic acids include DNA, RNA and analogs thereof, including peptide nucleic acids (PNA) and mixtures thereof. Nucleic acids can be single or two stranded. When referring to probes or primers, optionally labeled, with a detectable label, such as a fluorescent or radiolabel, single-stranded molecules are contemplated. Such molecules are typically of a length such that their targets are statistically unique or of low copy number (typically less than 5, generally less than 3) for probing or priming a library. Generally a probe or primer contains at least 14, 16 or 30 contiguous of sequence complementary to or identical to a gene of interest. Probes and primers can be 10, 20, 30, 50, 100 or more nucleic acids long.
- nucleic acid encoding a fragment or portion of an endotheliase refers to a nucleic acid encoding only the recited fragment or portion of endotheliase protein, and not the other contiguous portions of the endotheliase as a continuous sequence.
- heterologous nucleic acid is nucleic acid that, if it is DNA encodes RNA, or, if RNA, encodes proteins that generally are not normally produced in vivo by the cell in which it is expressed or that mediates or encodes mediators that alter expression of endogenous nucleic acid, such as DNA, by affecting transcription, translation, or other regulatable biochemical processes or that is located in a different locus from its normal locus.
- Heterologous nucleic acid is generally not endogenous to the cell into which it is introduced, but has been obtained from another cell or prepared synthetically. Generally, although not necessarily, such nucleic acid encodes RNA and proteins that are not normally produced by the cell in which it is now expressed.
- heterologous nucleic acid such as DNA
- foreign nucleic acid such as DNA
- Any nucleic acid, such as DNA, that one of skill in the art would recognize or consider as heterologous or foreign to the cell in which is expressed is herein encompassed by heterologous nucleic acid; heterologous nucleic acid includes exogenously added nucleic acid that is also expressed endogenously.
- heterologous nucleic acid examples include, but are not limited to, nucleic acid that encodes traceable marker proteins, such as a protein that confers drug resistance, nucleic acid that encodes therapeutically effective substances, such as anti-cancer agents, enzymes and hormones, and nucleic acid, such as DNA, that encodes other types of proteins, such as antibodies, and RNA, such as RNA interference (RNAi) or other double-stranded RNA, and antisense RNA.
- RNAi RNA interference
- Antibodies that are encoded by heterologous nucleic acid can be secreted or expressed on the surface of the cell in which the heterologous nucleic acid has been introduced.
- nucleic acid can be the the targeted agent, such as the therapeutic or diagnostic agent, in the conjugate.
- Nucleic acids include ds RNA use for RNA interference (RNAi) (see, e.g. Chuang et al. (2000) Proc. Natl. Acad. Sci. U.S.A. 97:4985) which is employed to inhibit the expression of a targeted gene by generating loss-of-function.
- RNAi RNA interference
- Methods relating to the use of RNAi to silence genes in organisms including, mammals, C. elegans , Drosophila and plants, and humans are known (see, e.g., Fire et al. (1998) Nature 391:806-811 Fire (1999) Trends Genet .
- dsRNA can interfere with accumulation of endogenous mRNA encoding a targeted gene product. Regions that include at least about 21 nucleotides and that are selective (i.e. whose target is unique) for the nucleic acid encoding a targeted gene product are used to prepare the RNAi.
- genetic therapy involves the transfer of heterologous nucleic acid, such as DNA, into certain cells, target cells, of a mammal, particularly a human, with a disorder or conditions for which such therapy is sought.
- the nucleic acid molecules are included in a conjugate linked via a cell surface protein cleavage site.
- the nucleic acid, such as DNA is introduced into the selected target cells in a manner such that the heterologous nucleic acid, such as DNA, is expressed and a therapeutic product encoded thereby is produced.
- the heterologous nucleic acid such as DNA
- Genetic therapy can also be used to deliver nucleic acid encoding a gene product that replaces a defective gene or supplements a gene product produced by the mammal or the cell in which it is introduced.
- the introduced nucleic acid can encode a therapeutic compound, such as a growth factor inhibitor thereof, or a tumor necrosis factor or inhibitor thereof, such as a receptor therefor, that is not normally produced in the mammalian host or that is not produced in therapeutically effective amounts or at a therapeutically useful time.
- the heterologous nucleic acid, such as DNA, encoding the therapeutic product can be modified prior to introduction into the cells of the afflicted host in order to enhance or otherwise alter the product or expression thereof. Genetic therapy can also involve delivery of an inhibitor or repressor or other modulator of gene expression, such dsRNA or antisense or other nucleic acid molecule.
- the conjugates herein can be used to deliver a product, such as a nucleic acid for gene therapy.
- a therapeutically effective product for gene therapy is a product that is encoded by heterologous nucleic acid, typically DNA, that, upon introduction of the nucleic acid into a host, a product is expressed that ameliorates or eliminates the symptoms, manifestations of an inherited or acquired disease or that cures the disease.
- heterologous nucleic acid typically DNA
- biologically active nucleic acid molecules such as RNAi and antisense.
- a sequence complementary to at least a portion of an RNA means a sequence having sufficient complementarily to be able to hybridize with the RNA, generally under moderate or high stringency conditions, forming a stable duplex; in the case of double-stranded SP antisense nucleic acids, a single strand of the duplex DNA (or dsRNA) can thus be tested, or triplex formation can be assayed.
- the ability to hybridize depends on the degree of complementarily and the length of the antisense nucleic acid.
- the longer the hybridizing nucleic acid the more base mismatches with a SP encoding RNA it can contain and still form a stable duplex (or triplex, as the case can be).
- One skilled in the art can ascertain a tolerable degree of mismatch by use of standard procedures to determine the melting point of the hybridized complex.
- Amino acid substitutions can be made or occur in any SPs and protease domains thereof. Amino acid substitutions include conservative substitutions, such as those set forth in Table 1, which do not eliminate proteolytic activity. As described herein, substitutions that alter properties of the proteins, such as removal of cleavage sites and other such sites are also contemplated; such substitutions are generally non-conservative, but can be readily effected by those of skill in the art.
- Suitable conservative substitutions of amino acids are known to those of skill in this art and can be made generally without altering the biological activity, for example enzymatic activity, of the resulting molecule. Also included within the definition, is the catalytically active fragment of an SP, particularly a single chain protease portion.
- substitutions are also permissible and can be determined empirically or in accord with known conservative substitutions.
- one or more amino acid residues within the sequence can be substituted by another amino acid of a similar polarity which acts as a functional equivalent, resulting in a silent alteration.
- Substitutes for an amino acid within the sequence can be selected from other members of the class to which the amino acid belongs.
- the nonpolar (hydrophobic) amino acids include alanine, leucine, isoleucine, valine, proline, phenylalanine, tryptophan and methionine.
- the polar neutral amino acids include glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine.
- the positively charged (basic) amino acids include arginine, lysine and histidine.
- the negatively charged (acidic) amino acids include aspartic acid and glutamic acid.
- amino acids which occur in the various amino acid sequences appearing herein, are identified according to their well-known, three-letter or one-letter abbreviations.
- nucleotides which occur in the various DNA fragments, are designated with the standard single-letter designations used routinely in the art.
- hR or hArg for homoarginine
- hY or hTyr for homotyrosine
- Cha for cyclohexylalanine
- Amf for 4-aminomethylphenylalanine
- DPL for 2-(4,6-dimethylpyrimidinyl)lysine
- (imidazolyl)K for N′-(2-imidazolyl)lysine
- Me2PO3-Y for 0-dimethylphosphotyrosine
- O—Me—Y for O-methyltyrosine
- TIC for tetrahydro-3-isoquinoline carboxylic acid
- MeL for 2-keto-3-amino-5-methylhexane
- DAP for 1,3-diaminopropane
- TFA trifluoroacetic acid
- AA for acetic acid.
- a splice variant refers to a variant produced by differential processing of a primary transcript of genomic DNA that results in more than one type of mRNA.
- a probe or primer based on a nucleotide sequence disclosed herein includes at least 10, 14, generally at least 16 or 30 or 100 contiguous sequence of nucleotides.
- antisense polynucleotides refer to synthetic sequences of nucleotide bases complementary to mRNA or the sense strand of double-stranded DNA. Admixture of sense and antisense polynucleotides under appropriate conditions leads to the binding of the two molecules, or hybridization. When these polynucleotides bind to (hybridize with) mRNA, inhibition of protein synthesis (translation) occurs. When these polynucleotides bind to double-stranded DNA, inhibition of RNA synthesis (transcription) occurs. The resulting inhibition of translation and/or transcription leads to an inhibition of the synthesis of the protein encoded by the sense strand.
- Antisense nucleic acid molecules typically contain a sufficient number of nucleotides to specifically bind to a target nucleic acid, generally at least 5 contiguous nucleotides, often at least 14 or 16 or 30 contiguous nucleotides or modified nucleotides complementary to the coding portion of a nucleic acid molecule that encodes a gene of interest, for example, nucleic acid encoding a single chain protease domain of an SP.
- an array refers to a collection of elements, such as antibodies, containing three or more members.
- An addressable array is one in which the members of the array are identifiable, typically by position on a solid phase support. Hence, in general the members of the array are immobilized on discrete identifiable loci on the surface of a solid phase.
- antibody refers to an immunoglobulin, whether natural or partially or wholly synthetically produced, including any derivative thereof that retains the specific binding ability of the antibody.
- antibody includes any protein having a binding domain that is homologous or substantially homologous to an immunoglobulin binding domain.
- Antibodies include members of any immunoglobulin claims, including IgG, IgM, IgA, IgD and IgE.
- antibody fragment refers to any derivative of an antibody that is less than full-length, retaining at least a portion of the full-length antibody's specific binding ability.
- antibody fragments include,but are not limited to, Fab, Fab′, F(ab) 2 , single-chain Fvs (scFV), FV, dsFV diabody and Fd fragments.
- the fragment can include multiple chains linked together, such as by disulfide bridges.
- An antibody fragment generally contains at least about 50 amino acids and typically at least 200 amino acids.
- an Fv antibody fragment is composed of one variable heavy domain (V H ) and one variable light domain linked by noncovalent interactions.
- a dsFV refers to an Fv with an engineered intermolecular disulfide bond, which stabilizes the V H -V L pair.
- an F(ab) 2 fragment is an antibody fragment that results from digestion of an immunoglobulin with pepsin at pH 4.0-4.5; it can be recombinantly expressed to produce the equivalent fragment.
- Fab fragments are antibody fragments that result from digestion of an immunoglobulin with papain; they can be recombinantly expressed to produce the equivalent fragment.
- scFVs refer to antibody fragments that contain a variable light chain (V L ) and variable heavy chain (V H ) covalently connected by a polypeptide linker in any order.
- the linker is of a length such that the two variable domains are bridged without substantial interference.
- exemplary linkers include, but are not limited to, (Gly-Ser) n residues, which can include ome Glu or Lys residues dispersed throughout, for example, to increase solubility.
- humanized antibodies refer to antibodies that are modified to include human sequences of amino acids so that administration to a human does not provoke an immune response.
- Methods for preparation of such antibodies are known.
- the encoding nucleic acid in the hybridoma or other prokaryotic or eukaryotic cell, such as an E. coli or a CHO cell, that expresses the monoclonal antibody is altered by recombinant nucleic acid techniques to express an antibody in which the amino acid composition of the non-variable region is based on human antibodies.
- Computer programs have been designed to identify such non-variable regions.
- diabodies are dimeric scFV; diabodies typically have shorter peptide linkers than scFvs, and they generally dimerize.
- production by recombinant means by using recombinant DNA methods means the use of the well known methods of molecular biology for expressing proteins encoded by cloned DNA.
- the term assessing is intended to include quantitative and qualitative determination in the sense of obtaining an absolute value for the activity of an SP, or a domain thereof, present in the sample, and also of obtaining an index, ratio, percentage, visual or other value indicative of the level of the activity. Assessment can be direct or indirect and the chemical species actually detected need not of course be the proteolysis product itself but can for example be a derivative thereof or some further substance.
- biological activity refers to the in vivo activities of a compound or physiological responses that result upon in vivo administration of a compound, composition or other mixture.
- Biological activity thus, encompasses therapeutic effects and pharmaceutical activity of such compounds, compositions and mixtures.
- Biological activities can be observed in in vitro systems designed to test or use such activities.
- a combination refers to any association between two or among more items.
- fluid refers to any composition that can flow. Fluids thus encompass compositions that are in the form of semi-solids, pastes, solutions, aqueous mixtures, gels, lotions, creams and other such compositions.
- an effective amount of a compound for treating a particular disease is an amount that is sufficient to ameliorate, or in some manner reduce the symptoms associated with the disease.
- Such amount can be administered as a single dosage or can be administered according to a regimen, whereby it is effective.
- the amount can cure the disease but, typically, is administered in order to ameliorate the symptoms of the disease. Repeated administration can be required to achieve the desired amelioration of symptoms.
- equivalent when referring to two sequences of nucleic acids, means that the two sequences in question encode the same sequence of amino acids or equivalent proteins.
- equivalent when equivalent is used in referring to two proteins or peptides, it means that the two proteins or peptides have substantially the same amino acid sequence with amino acid substitutions (see, e.g., Table 1, above) that do not substantially alter the activity or function of the protein or peptide (i.e, retain at least about 1% of the activity).
- equivalent refers to a property, the property does not need to be present to the same extent (e.g., two peptides can exhibit different rates of the same type of enzymatic activity), but the activities are generally substantially the same.
- Complementary when referring to two nucleotide sequences, means that the two sequences of nucleotides are capable of hybridizing, typically with less than 25%, often with less than 15%, or even less than 5% or with no mismatches between opposed nucleotides. Generally the two molecules hybridize under conditions of high stringency.
- a method for treating or preventing disease or disorder associated with undesired and/or uncontrolled angiogenesis means that the diseases or the symptoms associated with the undesired and/or uncontrolled angiogenesis are alleviated, reduced, ameliorated, prevented, placed in a state of remission, or maintained in a state of remission. It also means that the hallmarks of pathological angiogenesis are eliminated, reduced or prevented by the treatment.
- Non-limiting examples of the hallmarks of the pathological angiogenesis include uncontrolled degradation of the basement membrane and proximal extracellular matrix of the endothelial cells, migration, division, and organization of the endothelial cells into new functioning capillaries, and the persistence of such functioning capillaries.
- operatively linked or operationally associated refers to the functional relationship of DNA with regulatory and effector sequences of nucleotides, such as promoters, enhancers, transcriptional and translational stop sites, and other signal sequences.
- operative linkage of DNA to a promoter refers to the physical and functional relationship between the DNA and the promoter such that the transcription of such DNA is initiated from the promoter by an RNA polymerase that specifically recognizes, binds to and transcribes the DNA.
- a promoter region or promoter element refers to a segment of DNA or RNA that controls transcription of the DNA or RNA to which it is operatively linked.
- the promoter region includes specific sequences that are sufficient for RNA polymerase recognition, binding and transcription initiation. This portion of the promoter region is referred to as the promoter.
- the promoter region includes sequences that modulate this recognition, binding and transcription initiation activity of RNA polymerase. These sequences can be cis acting or can be responsive to trans acting factors. Promoters, depending upon the nature of the regulation, can be constitutive or regulated. Exemplary promoters contemplated for use in prokaryotes include the bacteriophage T7 and T3 promoters.
- sample refers to anything which can contain an analyte for which an analyte assay is desired.
- the sample can be a biological sample, such as a biological fluid or a biological tissue.
- biological fluids include urine, blood, plasma, serum, saliva, semen, stool, sputum, cerebral spinal fluid, tears, mucus, amniotic fluid or the like.
- Biological tissues are aggregates of cells, usually of a particular kind together with their intercellular substance that form one of the structural materials of a human, animal, plant, bacterial, fungal or viral structure, including connective, epithelium, muscle and nerve tissues. Examples of biological tissues also include organs, tumors, lymph nodes, arteries and individual cell(s).
- hybridize under conditions of a specified stringency is used to describe the stability of hybrids formed between two single-stranded DNA fragments and refers to the conditions of ionic strength and temperature at which such hybrids are washed, following annealing under conditions of stringency less than or equal to that of the washing step.
- high, medium and low stringency encompass the following conditions or equivalent conditions thereto:
- low stringency 1.0 ⁇ SSPE or SSC, 0.1% SDS, 50° C.
- Equivalent conditions refer to conditions that select for substantially the same percentage of mismatch in the resulting hybrids. Additions of ingredients, such as formamide, Ficoll, and Denhardt's solution affect parameters such as the temperature under which the hybridization should be conducted and the rate of the reaction. Thus, hybridization in 5 ⁇ SSC, in 20% formamide at 42° C. is substantially the same as the conditions recited above hybridization under conditions of low stringency.
- the recipes for SSPE, SSC and Denhardt's and the preparation of deionized formamide are described, for example, in Sambrook et al.
- substantially identical to a product means sufficiently similar so that the property of interest is sufficiently unchanged so that the substantially identical product can be used in place of the product.
- target cell refers to a cell that expresses a cell surface protease.
- a therapeutic agent As used herein, the terms a therapeutic agent, therapeutic regimen, radioprotectant, chemotherapeutic mean conventional drugs and drug therapies, including vaccines, which are known to those skilled in the art. Radiotherapeutic agents are well known in the art.
- vector refers to discrete elements that are used to introduce heterologous DNA into cells for expression and/or replication thereof.
- the vectors typically remain episomal, but can be designed to effect integration of a gene or portion thereof into a chromosome of the genome.
- vectors that are artificial chromosomes such as yeast artificial chromosomes and mammalian artificial chromosomes. Selection and use of such vehicles are well known to those of skill in the art.
- An expression vector includes vectors capable of expressing DNA that is operatively linked with regulatory sequences, such as promoter regions, that are capable of effecting expression of such DNA fragments.
- an expression vector refers to a recombinant DNA or RNA construct, such as a plasmid, a phage, recombinant virus or other vector that, upon introduction into an appropriate host cell, results in expression of the cloned DNA.
- Appropriate expression vectors are well known to those of skill in the art and include those that are replicable in eukaryotic cells and/or prokaryotic cells and those that remain episomal or those which integrate into the host cell genome.
- chemically stable means that the compound is stable enough to be formulated for pharmaceutical use. Such chemical stability is well known to those of skill in the art and can be determined by well known routine methods. Whether a given compound is chemically stable enough to be formulated for pharmaceutical use depends on a number of factors including, but not limited to, the type of formulation and route of administration desired, the disease to be treated, and the method of preparing the pharmaceutical formulation.
- a “functional equivalent” of a side chain of an amino acid is a group or moiety that functions in substantially the same way as the naturally occurring side chain to achieve substantially the same result (e.g., a substrate for a cell surface protease).
- side chain of arginine include, but are not limited to, homoarginine, guanidinoaminopropyl, guanidinoaminoethyl, (Me) 2 arginine side chain, (Et) 2 arginine side chain, (4-aminomethyl)phenylmethyl, 4-amidinophenylmethyl, 4-guanidinophenyl-methyl, or a conformationally constrained arginine side chain analog such as:
- x is 0 or 1 (see, e.g., Webb et al. (1991) J. Org. Chem . 56:3009), or a conformationally constrained arginine side chain analog such as:
- d is an integer from 0 to 5, or 1 to 3; and W is N or CH; or a mono- or di-substituted N-alkyl derivative of the above groups, where alkyl is, in certain embodiments, lower alkyl, such as methyl.
- pharmaceutically acceptable derivatives of a compound include salts, esters, enol ethers, enol esters, acids, bases, solvates, hydrates or prodrugs thereof.
- Such derivatives can be readily prepared by those of skill in this art using known methods for such derivatization.
- the compounds produced can be administered to animals or humans without substantial toxic effects and either are pharmaceutically active or are prodrugs.
- salts include, but are not limited to, amine salts, such as but not limited to N,N′-dibenzylethylenediamine, chloroprocaine, choline, ammonia, diethanolamine and other hydroxyalkylamines, ethylenediamine, N-methylglucamine, procaine, N-benzylphenethylamine, 1-para-chlorobenzyl-2-pyrrolidin-1′-ylmethylbenzimidazole, diethylamine and other alkylamines, piperazine and tris(hydroxymethyl)aminomethane; alkali metal salts, such as but not limited to lithium, potassium and sodium; alkali earth metal salts, such as but not limited to barium, calcium and magnesium; transition metal salts, such as but not limited to zinc; and other metal salts, such as but not limited to sodium hydrogen phosphate and disodium phosphate; and also including, but not limited to, salts of mineral acids, such as but not limited to hydrochlorides and
- esters include, but are not limited to, alkyl, alkenyl, alkynyl, aryl, heteroaryl, aralkyl, heteroaralkyl, cycloalkyl and heterocyclyl esters of acidic groups, including, but not limited to, carboxylic acids, phosphoric acids, phosphinic acids, sulfonic acids, sulfinic acids and boronic acids.
- Pharmaceutically acceptable enol ethers include, but are not limited to, derivatives of formula C ⁇ C(OR) where R is hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, aralkyl, heteroaralkyl, cycloalkyl or heterocyclyl.
- enol esters include, but are not limited to, derivatives of formula C ⁇ C(OC(O)R) where R is hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, aralkyl, heteroaralkyl, cycloalkyl ar heterocyclyl.
- Pharmaceutically acceptable solvates and hydrates are complexes of a compound with one or more solvent or water molecule, generally 1 to about 100, typically 1 to about 10, such as 1 to about 2, 3 or 4, solvent or water molecules.
- treatment means any manner in which one or more of the symptoms of a condition, disorder or disease are ameliorated or otherwise beneficially altered. Treatment also encompasses any pharmaceutical use of the compositions herein, such as use for treating cancer.
- amelioration of the symptoms of a particular disorder by administration of a particular pharmaceutical composition refers to any lessening, whether permanent or temporary, lasting or transient that can be attributed to or associated with administration of the composition.
- a prodrug is a compound that, upon in vivo administration, is metabolized or otherwise converted to the biologically, pharmaceutically or therapeutically active form of the compound.
- the pharmaceutically active compound is modified such that the active compound is regenerated by metabolic processes.
- the prodrug can be designed to alter the metabolic stability or the transport characteristics of a drug, to mask side effects or toxicity, to improve the flavor of a drug or to alter other characteristics or properties of a drug.
- the conjugates provided herein can contain chiral centers. Such chiral centers can be of either the (R) or (S) configuration, or can be a mixture thereof.
- the compounds provided herein can be enantiomerically pure, or be stereoisomeric or diastereomeric mixtures.
- amino acid residues such residues can be of either the L- or D-form.
- the configuration for naturally occurring amino acid residues is generally L. When not specified the residue is the L form.
- the chiral centers of the compounds provided herein can undergo epimerization in vivo. As such, one of skill in the art will recognize that administration of a compound in its (R) form is equivalent, for compounds that undergo epimerization in vivo, to administration of the compound in its (S) form.
- the conjugates provided herein are prodrugs because they include a therapeutic agent in an inactive form that is ultimately converted to an active form at the targeted cell or tissue or in the environment thereof.
- a biologically, pharmaceutically or therapeutically active form of a compound is released, or, a derivative that can be further metabolized into a biologically, pharmaceutically or therapeutically active form of a compound.
- substantially pure means sufficiently homogeneous to appear free of readily detectable impurities as determined by standard methods of analysis, such as thin layer chromatography (TLC), gel electrophoresis, high performance liquid chromatography (HPLC) and mass spectrometry (MS), used by those of skill in the art to assess such purity, or sufficiently pure such that further purification would not alter the physical and chemical properties, such as enzymatic and biological activities, of the substance for its intended purpose.
- TLC thin layer chromatography
- HPLC high performance liquid chromatography
- MS mass spectrometry
- a peptidic substrate includes peptides and molecules, such as peptide mimetics and peptides that include peptide bond surrogates.
- Amino acids (or amino acid surrogates) that form the scissile bond are assigned the number 1, adjacent residues the number 2, and so on, counting away from the scissile bond.
- Each specific subsite of the substrate therefore, is uniquely identified by a number and the designation as primed or unprimed.
- a surrogate of a peptide bond is a divalent group that possesses similar steric and/or electronic characteristics to —C(O)NH—.
- Peptide bond surrogates include, but are not limited to, alkene isosteres (—CR ⁇ CR—), particularly (E)-alkene isosteres of formula —CH ⁇ CH—, hydroxyethylene isosteres (—CH(OH)CH 2 —), enamine isosteres (—C( ⁇ CRR)NH—), aminoalcohol isosteres (—CH(OH)CH 2 NH—), difluoroketone isosteres (—C(O)CF 2 —), retroinverso compounds (—NHC(O)—), divalent heterocyclyl or heteroaryl groups, and cyclopropyl isosteres such as:
- alkyl, alkenyl and alkynyl carbon chains contain from 1 to 20 carbons, generally 1 to 16 carbons, and are straight or branched.
- Alkenyl carbon chains of from 2 to 20 carbons typically contain 1 to 8 double bonds, and the alkenyl carbon chains of 2 to 16 carbons and typically contain 1 to 5 double bonds.
- Alkynyl carbon chains of from 2 to 20 carbons typically contain 1 to 8 triple bonds, and the alkynyl carbon chains of 2 to 16 carbons and generally contain 1 to 5 triple bonds.
- alkyl, alkenyl and alkynyl groups herein include, but are not limited to, methyl, ethyl, propyl, isopropyl, isobutyl, n-butyl, sec-butyl, tert-butyl, isopentyl, neopentyl, tert-pentyl and isohexyl.
- the alkyl, alkenyl and alkynyl groups optionally can be substituted, with one or more groups, generally alkyl group substituents that are the same or different.
- lower alkyl, lower alkenyl, and lower alkynyl refer to carbon chains having less than about 6 carbons.
- alk(en)(yn)yl refers to an alkyl group containing at least one double bond and at least one triple bond.
- cycloalkyl refers to a saturated mono- or multi-cyclic ring system, typically 3 to 10 carbon atoms, such as, for example, 3 to 6 carbon atoms; cycloalkenyl and cycloalkynyl refer to mono- or multicyclic ring systems that respectively include at least one double bond and at least one triple bond. Cycloalkenyl and cycloalkynyl groups contain, for example, 3 to 10 carbon atoms, with cycloalkenyl groups generally containing 4 to 7 carbon atoms and cycloalkynyl groups that contain, for example 8 to 10 carbon atoms.
- ring systems of the cycloalkyl, cycloalkenyl and cycloalkynyl groups can be composed of one ring or two or more rings which can be joined together in a fused, bridged or spiro-connected fashion, and optionally can be substituted with one or more alkyl group substituents.
- Cycloalk(en)(yn)yl refers to a cycloalkyl group containing at least one double bond and at least one triple bond.
- substituted alkyl refers to alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl and cycloalkynyl groups, respectively, that are substituted with one or more substituents, in certain embodiments one to three substituents, independently selected from alkyl, halo, haloalkyl, such as halo lower alkyl, pseudohalo, aryl, amino, dialkylamino, nitro, cyano, azido, alkylsulfinyl, alkylsulfonyl, alkylcarbonylamino, alkoxycarbonylamino, aminoimino, hydroxy, alk
- aryl refers to cyclic groups containing from 6 to 19 carbon atoms.
- Aryl groups include, but are not limited to groups, such as fluorenyl, substituted fluorenyl, phenyl, substituted phenyl, naphthyl and substituted naphthyl.
- aryl also refers to aryl-containing groups, including, but not limited to, aryloxy, arylthio, arylcarbonyl and arylamino groups.
- heteroaryl refers to a monocyclic or multicyclic aromatic ring system, generally about 5 to about 15 members where one or more, such as 1 to 3 of the atoms in the ring system is a heteroatom, that is, an element other than carbon, for example, nitrogen, oxygen and sulfur atoms.
- the heteroaryl group optionally can be fused to a benzene ring.
- heteroaryl groups include, for example, furyl, imidazolyl, pyrrolidinyl, pyrimidinyl, tetrazolyl, thienyl, pyridyl, pyrrolyl, N-methylpyrrolyl, quinolinyl and isoquinolinyl, with pyridyl, thienyl and quinolinyl as examples thereof.
- heteroaryl also refers to heteroaryl-containing groups, including, but not limited to, heteroaryloxy, heteroarylthio, heteroarylcarbonyl and heteroarylamino.
- heterocyclyl refers to a monocyclic or multicyclic non-aromatic ring system, such as systems of 3 to 10 members, for exmaple 4 to 7 members or 5 to 6 members, where one or more, such as 1 to 3 of the atoms in the ring system is a heteroatom, that is, an element other than carbon, for example, nitrogen, oxygen and/or sulfur atoms.
- substituted aryl refers to aryl, heteroaryl and heterocyclyl groups, respectively, that are substituted with one or more substituents, in certain embodiments one to three substituents, independently selected from alkyl, cycloalkyl, cycloalkylalkyl, aryl, heteroaryl optionally substituted with 1 or more, such as 1 to 3, substituents selected from halo, halo alkyl and alkyl, aralkyl, heteroaralkyl, alkenyl containing 1 to 2 double bonds, alkynyl containing 1 to 2 triple bonds, alk(en)(yn)yl groups, halo, pseudohalo, cyano, hydroxy, haloalkyl and polyhaloalkyl, such as halo lower alkyl, especially trifluoromethyl, formyl, alkylcarbonyl,
- aralkyl refers to an alkyl group in which one of the hydrogen atoms of the alkyl is replaced by an aryl group.
- heteroarylkyl refers to an alkyl group in which one of the hydrogen atoms of the alkyl is replaced by a heteroaryl group.
- alkyl refers to saturated carbon chains that contain one or more carbons; the chains can be straight or branched or include cyclic portions or be cyclic.
- haloalkyl can include one or more of the same or different halogens.
- C 1-3 alkoxyphenyl can include one or more of the same or different alkoxy groups containing one, two or three carbons.
- halo refers to F, Cl, Br or I.
- pseudohalides are compounds that behave substantially similar to halides. Such compounds can be used in the same manner and treated in the same manner as halides (X ⁇ , in which X is a halogen, such as Cl or Br).
- Pseudohalides include, but are not limited to, cyanide, cyanate, thiocyanate, selenocyanate, trifluoromethoxy, difluoromethoxy, dichloromethoxy and azide.
- haloalkyl refers to a lower alkyl radical in which one or more of the hydrogen atoms are replaced by halogen.
- groups include, but not limited to, chloromethyl, trifluoromethyl, 1-chloro-2-fluoroethyl and the like.
- haloalkoxy refers to RO— in which R is a haloalkyl group.
- sulfinyl or “thionyl” refers to —S(O)—.
- sulfonyl or “sulfuryl” refers to —S(O) 2 —.
- sulfo refers to —S(O) 2 O—.
- Carboxy refers to a divalent radical, —C(O)O—.
- aminocarbonyl refers to —C(O)NH 2 .
- alkylaminocarbonyl refers to —C(O)NHR in which R is hydrogen or alkyl, such as, for example, lower alkyl.
- dialkylaminocarbonyl refers to —C(O)NR′R in which R′ and R are independently selected from hydrogen or alkyl, such as, for example, lower alkyl; “carboxamide” refers to groups of formula —NR′COR.
- diarylaminocarbonyl refers to —C(O)NRR′ in which R and R′ are independently selected from aryl, such as lower aryl, for example, phenyl.
- aralkylaminocarbonyl refers to —C(O)NRR′ in which one of R and R′ is aryl, such as, lower aryl, for example, phenyl, and the other of R and R′ is alkyl, such as, for example, lower alkyl.
- arylaminocarbonyl refers to —C(O)NHR in which R is aryl, such as lower aryl, for example, phenyl.
- hydroxycarbonyl refers to —COOH
- alkoxycarbonyl refers to —C(O)OR in which R is alkyl, such as lower alkyl.
- aryloxycarbonyl refers to —C(O)OR in which R is aryl, such lower aryl, for example phenyl.
- alkoxy and “alkylthio” refer to RO— and RS—, in which R is alkyl, such as, for example, lower alkyl.
- aryloxy and “arylthio” refer to RO— and RS—, in which R is aryl, such lower aryl, for example, phenyl.
- alkylene refers to a straight, branched or cyclic, such as, for example, straight or branched, divalent aliphatic hydrocarbon group, for example, having from 1 to about 20 carbon atoms such as 1 to 12 carbons, and for exmaple, is lower alkylene. There optionally can be inserted along the alkylene group one or more oxygen, sulphur or substituted or unsubstituted nitrogen atoms, where the nitrogen substituent is alkyl as previously described.
- Exemplary alkylene groups include methylene (—CH 2 —), ethylene (—CH 2 CH 2 —), propylene (—(CH 2 ) 3 —), cyclohexylene (—C 6 H 10 —), methylenedioxy (—O—CH 2 —O—) and ethylenedioxy (—O—(CH 2 ) 2 —O—).
- the term “lower alkylene” refers to alkylene groups having 1 to 6 carbons.
- Exemplary alkylene groups are lower alkylene, such as, for example, alkylene of 1 to 3 carbon atoms.
- alkenylene refers to a straight, branched or cyclic, typically straight or branched, divalent aliphatic hydrocarbon group, such as, for example, having from 2 to about 20 carbon atoms and at least one double bond, generally 1 to 12 carbons, and is for example, lower alkenylene. There optionally can be inserted along the alkenylene group one or more oxygen, sulphur or substituted or unsubstituted nitrogen atoms, where the nitrogen substituent is alkyl as previously described. Exemplary alkenylene groups include —CH ⁇ CH—CH ⁇ CH— and —CH ⁇ CH ⁇ CH 2 —.
- the term “lower alkenylene” refers to alkenylene groups having 2 to 6 carbons. Examplary alkenylene groups are lower alkenylene, such as, for example, alkenylene of 3 to 4 carbon atoms.
- alkynylene refers to a straight, branched or cyclic, generally straight or branched, divalent aliphatic hydrocarbon group, such those having from 2 to about 20 carbon atoms and at least one triple bond, generally 1 to 12 carbons, such as, for example, lower alkynylene. There optionally can be inserted along the alkynylene group one or more oxygen, sulphur or substituted or unsubstituted nitrogen atoms, where the nitrogen substituent is alkyl as previously described. Exemplary alkynylene groups include —C ⁇ C—C ⁇ C—, —C ⁇ C— and —C ⁇ C—CH 2 —.
- the term “lower alkynylene” refers to alkynylene groups having 2 to 6 carbons. Exemplary alkynylene groups are lower alkynylene, such as, for example, alkynylene of 3 to 4 carbon atoms.
- alk(en)(yn)ylene refers to a straight, branched or cyclic, generally straight or branched, divalent aliphatic hydrocarbon group, having, for example, from 2 to about 20 carbon atoms and at least one triple bond, and at least one double bond; typically 1 to 12 carbons, such as, for example, lower alk(en)(yn)ylene.
- Exemplary alk(en)(yn)ylene groups include —C ⁇ C—(CH 2 ) n —C ⁇ C—, where n is 1 or 2.
- the term “lower alk(en)(yn)ylene” refers to alk(en)(yn)ylene groups having up to 6 carbons.
- Exemplary alk(en)(yn)ylene groups are lower alk(en)(yn)ylene, such as, for example, alk(en)(yn)ylene of 4 carbon atoms.
- cycloalkylene refers to a divalent saturated mono- or multicyclic ring system, generally 3 to 10 carbon atoms, such as 3 to 6 carbon atoms; cycloalkenylene and cycloalkynylene refer to divalent mono- or multicyclic ring systems that respectively include at least one double bond and at least one triple bond. Cycloalkenylene and cycloalkynylene groups can contain 3 to 10 carbon atoms, with, for example, cycloalkenylene groups containing 4 to 7 carbon atoms and cycloalkynylene groups containing 8 to 10 carbon atoms.
- ring systems of the cycloalkylene, cycloalkenylene and cycloalkynylene groups can be composed of one ring or two or more rings that can be joined together in a fused, bridged or spiro-connected fashion.
- Cycloalk(en)(yn)ylene refers to a cycloalkylene group containing at least one double bond and at least one triple bond.
- substituted alkylene refers to alkylene, alkenylene, alkynylene, cycloalkylene, cycloalkenylene and cycloalkynylene groups, respectively, that are substituted with one or more substituents, in certain embodiments one to three substituents, independently selected from halo, haloalkyl, such as, for example, halo lower alkyl, aryl, hydroxy, alkoxy, aryloxy, alkyloxy, alkylthio, arylthio, aralkyloxy, aralkylthio, carboxy alkoxycarbonyl, oxo and cycloalkyl.
- arylene refers to a monocyclic or polycyclic, such as monocyclic, divalent aromatic group, for example, having from 5 to about 20 carbon atoms and at least one aromatic ring, such as 5 to 12 carbons, and, is, for example, lower arylene. There optionally can be inserted around the arylene group one or more oxygen, sulphur or substituted or unsubstituted nitrogen atoms, where the nitrogen substituent is alkyl as previously described. Exemplary arylene groups include 1,2-, 1,3- and 1,4-phenylene.
- the term “lower arylene” refers to arylene groups having 5 or 6 carbons. Exemplary arylene groups are lower arylene.
- heteroarylene refers to a divalent monocyclic or multicyclic aromatic ring system, such as of about 5 to about 15 members where one or more, typically, for example, 1 to 3 of the atoms in the ring system is a heteroatom, that is, an element other than carbon, for example, nitrogen, oxygen and/or sulfur atom(s).
- heterocyclylene refers to a divalent monocyclic or multicyclic non-aromatic ring system, generally of 3 to 10 members, such as, for example, 4 to 7 members or 5 to 6 members, where one or more, such as, for example, 1 to 3 of the atoms in the ring system is a heteroatom, that is, an element other than carbon, for example, nitrogen, oxygen and/or sulfur atom(s).
- substituted arylene refers to arylene, heteroarylene and heterocyclylene groups, respectively, that are substituted with one or more substituents, in certain embodiments one to three substituents, independently selected from alkyl, cycloalkyl, cycloalkylalkyl, aryl, heteroaryl optionally substituted with 1 or more, such as 1 to 3, substituents selected from halo, halo alkyl and alkyl, aralkyl, heteroaralkyl, alkenyl containing 1 to 2 double bonds, alkynyl containing 1 to 2 triple bonds, alk(en)(yn)yl groups, halo, pseudohalo, cyano, hydroxy, haloalkyl and polyhaloalkyl, such as, halo lower alkyl, for example trifluoromethyl, formyl, alkylcarbonyl
- alkylidene refers to a divalent group, such as ⁇ CR′R′′, which is attached to one atom of another group, forming a double bond.
- exemplary alkylidene groups are methylidene ( ⁇ CH 2 ) and ethylidene ( ⁇ CHCH 3 ).
- aralkylidene refers to an alkylidene group in which either R′ or R′′ is an aryl group.
- Cycloalkylidene are those where R′ and R′′ are linked to form a carbocyclic ring.
- Heterocyclylidene are those where at least one of R′ and R′′ contain a heteroatom in the chain, and R′ and R′′ are linked to form a heterocyclic ring.
- amido refers to the divalent group —C(O)NH—.
- Thioamido refers to the divalent group —C(S)NH—.
- Oxyamido refers to the divalent group —OC(O)NH—.
- Thiaamido refers to the divalent group —SC(O)NH—.
- Dithiaamido refers to the divalent group —SC(S)NH—.
- Ureido refers to the divalent group —HNC(O)NH—.
- Thioureido refers to the divalent group —HNC(S)NH—.
- “semicarbazide” refers to —NHC(O)NHNH—. “Carbazate” refers to the divalent group —OC(O)NHNH—. “Isothiocarbazate” refers to the divalent group —SC(O)NHNH—. “Thiocarbazate” refers to the divalent group —OC(S)NHNH—. “Sulfonylhydrazide” refers to the group —SO 2 NHNH—. “Hydrazide” refers to the divalent group —C(O)NHNH—. “Azo” refers to the divalent group —N ⁇ N—. “Hydrazinyl” refers to the divalent group —NH—NH—.
- amino acid refers to ⁇ -amino acids which are racemic, or of either the D- or L-configuration.
- the designation “d” preceding an amino acid designation refers to the D-isomer of the amino acid.
- the designation “dI” preceding an amino acid designation refers to a mixture of the L- and D-isomers of the amino acid.
- HHT and CHT refer to hexahydrotyrosyl (also known as cyclohexyltyrosyl or p-hydroxycyclohexylalanyl), CHA is cyclohexylalanyl, Pyr and pyroGlu refer to pyroglutamic acid, Pip is pipecolinic acid, Sar is sarcosine, nLeu and Nle are norleucine, nVal is norvaline, Aib is 2-aminoisobutyric acid, Quat is (R)-Glu( ⁇ -(3-amidinobenzyl)), and Abu and But are 2-aminobutyric acid.
- PEG represents a polyethylene glycol containing substituent having the designated number of ethyleneoxy subunits.
- PEG(2) represents:
- cyclic moieties and heteroatom-containing cyclic moieties so defined include, but are not limited to:
- hydroxylated represents substitution on a substitutable carbon of the ring system being so described by a hydroxyl moiety.
- polyhydroxylated represents substitution on two or more substitutable carbons of the ring system being so described by 2, 3 or 4 hydroxyl moieties.
- cotininyl represents the following structure:
- 1-methylHis or (1Me)H refers to the structure:
- 3-methylHis or (3Me)H refers to the structure:
- Quat 2 refers to:
- Quat 3 refers to:
- Quat 4 refers to:
- Quat 5 refers to:
- the conjugates herein are designed to target proteases that are located on cell surfaces, particularly tumor cells and cells involved in tumorigenic processes and angiogenesis and other proliferative processes.
- the conjugates described in detail below, contain a peptidic substrate for a selected targeted cell surface protease linked, either directly or via a linker, to a therapeutic agent, typically a cytotoxic agent, which is substantially inactive when in the conjugate.
- a therapeutic agent typically a cytotoxic agent, which is substantially inactive when in the conjugate.
- the therapeutic agent is released in a form that is active or that can be activated in the vicinity of the targeted cell or tissue to which it is delivered. As a result, active therapeutic agent accumulates at the targeted cells or tissue or in the targeted cells.
- the targeted protease is selected by identifying a protease that is located on a cell or tissue (or associated therewith) that is involved in the disease process or serendipitously present in the locale of cells or tissues involved in the disease or disease process, and, generally, is not located at all or present or active at lower levels, generally substantially lower levels, or exhibits altered activity or specificity, on many, if not all, other cells or tissues.
- the variety and numbers of non-targeted cells or tissues that expresss the active protease varies for particular proteases and diseases intended for treatment. Those of skill in the art will select a target based upon the disease, targeted agents and tolerable or acceptable levels of side-effects. The goal is to achieve enhanced therapeutic index compared with administration of the targeted agent by itself.
- the targeted protease may or may not be involved in the disease process and its expression can be serendiptous; for purposes herein its particular role or lack thereof is not important; it is the fact that it is active in the locale of targeted tissues or cells that is important.
- many of the cell surface proteases of interest herein are expressed or active on tumor cells or cells involved in the tumorigenic processes. Any method known to one of skill in the art for determining or detecting a tissue or cell expression profile can be used.
- RNA blots composed of RNA from numerous tissues (e.g., a multiple tissue expression (MTE) array available from CLONTECH, Palo Alto, Calif.), can be screened with probes based upon the nucleic acid sequence of the protease of interest to identify cells that express the protease.
- Northern analysis of the blots to test for expression also can be used.
- MTSPs type II membrane-bound serine proteases
- proteases that are located at the cell surface by virtue of a specific interaction with a cell surface protein.
- Urokinase plasminogen activator (u-PA) bound to urokinase plasminogen activator receptor (u-PAR) is exemplary of such proteases.
- Nucleic acid sequence information and expression profiles of exemplary MTSPs and endotheliases are as follows (see, also EXAMPLE 6).
- Cell surface proteolysis is a mechanism for the generation of biologically active proteins that mediate a variety of cellular functions. These membrane-anchored proteins, include a disintegrin-like and metalloproteinase (ADAM) and membrane-type matrix metalloproteinase (MT-MMP).
- ADAM disintegrin-like and metalloproteinase
- MT-MMP membrane-type matrix metalloproteinase
- serine proteases have been implicated in neoplastic disease progression. Most serine proteases, which are either secreted enzymes or are sequestered in cytoplasmic storage organelles, have roles in blood coagulation, wound healing, digestion, immune responses and tumor invasion and metastasis.
- Transmembrane serine proteases appear to be involved in the etiology and pathogenesis of tumors. These enzymes are expressed in certain cancerous and tumor cells and in other cells associated with other proliferative disorders and other disease states, such as in inflammatory cells and and can be tissue or organ-specific. In mammals, more than 20 members of the family are known (see, Hooper et al. (2001) J. Biol. Chem . 276:857-860, see, also U.S. application Ser. No. 09/776,191, filed Feb. 2, 2001 and International PCT application No. PCT/US01/03471; see, also U.S. provisional application Serial Nos.
- accession nos. AF133086/AF118224, AF04280022 Takeuchi et al. (1999) Proc. Natl. Acad. Sci. U.S.A . 96:11054-1161; Lin et al. (1999) J. Biol. Chem . 274:18231-18236; Takeuchi et al. (2000) J. Biol. Chem . 275:26333-26342; and Kim et al. (1999) Immunogenetics 49:420-429); hepsin (see, accession nos. M18930, AF030065, X70900; Leytus et al. (1988) Biochem .
- proteases including transmembrane serine proteases, have been implicated in processes involved in neoplastic development and progression. While the precise role of these proteases has not been elaborated, serine proteases and inhibitors thereof are involved in the control of many intra- and extracellular physiological processes, including degradative actions in cancer cell invasion, metastatic spread, and neovascularization of tumors, that are involved in tumor progression. It is believed that proteases are involved in the degradation of extracellular matrix (ECM) and contribute to tissue remodeling, and are necessary for cancer invasion and metastasis. The activity and/or expression of some proteases have been shown to correlate with tumor progression and development, and also are shown to be active in specific cell types.
- ECM extracellular matrix
- MTSP1 membrane-type serine protease MTSP1 (also called matriptase; see SEQ ID Nos. 1 and 2 from U.S. Pat. No. 5,972,616; and GenBank Accession No. AF118224; (1999) J. Biol. Chem . 274:18231-18236; U.S. Pat. No. 5,792,616; see, also Takeuchi (1999) Proc. Natl. Acad. Sci. U.S.A . 96:11054-1161) that is expressed in epithelial cancer and normal tissue (Takeucuhi et al. (1999) Proc. Natl. Acad. Sci. USA 96:11054-61) has been identified.
- Matriptase also is expressed in a variety of epithelial tissues with high levels of activity and/or expression in the human gastrointestinal tract and the prostate.
- Hepsin a cell surface serine protease identified in hepatoma cells, is overexpressed in ovarian cancer (Tanimoto et al. (1997) Cancer Res ., 57:2884-7).
- the hepsin transcript appears to be abundant in carcinoma tissue and is almost never expressed in normal adult tissue, including normal ovary. It has been suggested that hepsin is frequently overexpressed in ovarian tumors and therefore can be a candidate protease in the invasive process and growth capacity of ovarian tumor cells.
- NES1 normal epithelial cell-specific 1
- Each MTSP has a characteristic tissue expression profile; the MTSPs in particular, although not exclusively expressed or activated in tumors, exhibit characteristic tumor tissue expression or activation profiles.
- MTSPs can have different activity in a tumor cell from a non-tumor cell by virtue of a change in a substrate or cofactor therefor or other factor that would alter functional activity of the MTSP.
- each can serve as a diagnostic marker for particular tumors, by virtue of a level of activity and/or expression or function in a subject (i.e. a mammal, particularly a human) with neoplastic disease, compared to a subject or subjects that do not have the neoplastic disease.
- detection of activity (and/or expression) in a particular tissue can be indicative of neoplastic disease.
- they can serve as therapeutic targets, such as by administration of modulators of the activity thereof, or, as by administration of a prodrug specifically activated by one of the MTSPs.
- Each or any of the MTSPs can exhibit activity or expression levels or substrate specificities that differ in tumor cells from the levels in normal cells.
- tumor cells include, but are not limited to, colon, lung, prostate, breast, esophagous, pancreas, cervic, uterus, endometrium, and other solid tumors and in blood and lymphatic tumors.
- conjugates provided herein can be designed by selection of substrate specificity for treatment of any of such tumors and neoplastic conditions.
- tissue and gene profiles of some exemplary MTSPs. These profiles are not intended to define the full scope of expression or activation of these MTPSs, but demonstrate that MTSPs are expressed in tumors, and, hence there expression or activation or substrate specificity on the surface of tumor cells can be exploited in the methods herein and conjugates, designed in accord with the methods herein and as exemplified herein, that are cleaved by one or more of these MTSPs can be prepared and employed for treatment of neoplastic or other diseases or conditions or to target to cells that express these proteins on there surfaces.
- MTSP1 (also called matriptase) is a trypsin-like serine protease with broad spectrum cleavage activity and two potential regulatory modules. It was named “matriptase” based on its ability to degrade the extra-cellular matrix and its trypsin-like activity.
- MTSP1 When isolated from breast cancer cells (or T-47D cell conditioned medium), MTSP1 has been reported to be primarily in an uncomplexed form. MTSP1 has been isolated from human milk; when isolated from human milk, it was reported to be in one of two complexed forms, 95 kDa (the predominant form) and 110 kDa; uncomplexed MTSP1 was not detected (Liu, et al.
- MTSP1 exists as an uncomplexed protease when in its active state.
- HAI-1 hepatocyte growth factor inhibitor-1
- Kunitz-type serine protease inhibitor having activity against trypsin-like serine proteases.
- Nucleic acids encoding the protein designed matriptase were cloned from T-47D human breast cancer cell-conditioned medium (Lin et al. (1999) J. Biol. Chem . 274:18231-18236). Upon analysis of the cDNA, it was determined that the full length protease has 683 amino acids and contains three main structural regions: a serine protease domain near the carboxyl-terminal region, four tandem low-density lipoprotein receptor domains, and two tandem complement subcomponents C1r and C1s (see SEQ ID No. 1). Studies to identify additional serine proteases made by cancer cells were done using PC-3 cells.
- MT-SP1 serine protease termed “MT-SP1” (MTSP1) by the authors, reported to be a transmembrane protease was cloned (Takeuchi et al. (1999) Proc. Natl. Acad. Sci. U.S.A . 96:11054-11061). It was subsequently found that originally identified matriptase sequence is included in the translated sequence of the cDNA that encodes MTSP1. The nucleic acid encoding the protein originally designated matriptase is a partial MTSP1 clone that lacks 516 of the coding nucleotides (Takeuchi, et al., J. Biol.
- matriptase is a variant form of MTSP1.
- MTSP1 demonstrates trypsin-like protease activity and is a Type II transmembrane protein with an extracellular protease domain.
- PAR2 protease-activated receptor 2
- pro-HGF pro-hepatacyte growth factor
- sc-uPA single-chain urokinase-type plasminogen activator
- PAR2 functions in inflammation, cytoprotection and/or cell adhesion, while sc-uPa functions in tumor cell invasion and metastasis.
- HGF serves a growth and pro-angiogenic factor.
- SEQ ID Nos 1 and 2 An exemplary nucleotide sequence encoding a human MTSP1 is set forth in SEQ ID Nos 1 and 2. As previously noted SEQ ID No. 1 sets for an MTSP1-encoding nucleic acid sequence. This sequence is the longer version and includes the protease domain, which is common to both variants.
- MTSP1 is expressed in breast, prostate and colorectal tumors. Hence conjugates with substrates therefor can be used for treatment of such tumors.
- the MTSP3 transcript was detected in lung carcinoma (LX-1), colon adenocarcinoma (CX-1), colon adenocarcinoma (GI-112) and ovarian carcinoma (GI-102). No apparent signal was detected in another form of lung carcinoma (GI-117), breast carcinoma (GI-101), pancreatic adenocarcinoma (GI-103) and prostatic adenocarcinoma (PC3).
- RNA blot composed of 76 different human tissues (catalog number 7775-1; human multiple tissue expression (MTE) array; CLONTECH). As in the northern analysis of gel blot, a very strong signal was observed in the liver.
- the MTSP4 transcript was detected in breast carcinoma (GI-101), lung carcinoma (LX-1), colon adenocarcinoma (GI-112) and pancreatic adenocarcinoma (GI-103). No apparent signal was detected in another form of lung carcinoma (GI-117), colon adenocarcinoma (CX-1), ovarian carcinoma (GI-102). and prostatic adenocarcinoma (PC3).
- the MTSP4 transcript was also detected in LNCaP and PC-3 prostate cancer cell lines as well as in HT-1080 human fibrosarcoma cell line.
- MTSP6 is expressed at high levels in the colon. It also is expressd in the, stomach, trachea, mammary gland, thyroid gland, salivary gland, pituitary gland and pancreas. It is expressed at lower levels in other tissues (see EXAMPLE 6).
- MTSP6 also is expressed in several tumor cell lines including HeLa S3>colorectal adenocarcinoma (SW480)>leukemia MOLT-4>leukemia K-562.
- the MTSP6 transcript was strongly detected in lung carcinoma (LX-1), moderately detected in pancreatic adenocarcinoma (GI-103), weakly detected in ovarian carcinoma (GI-102); and weakly detected in colon adenocarcinoma (GI-112 and CX-1), breast carcinoma (GI-101), lung carcinoma (GI-117) and prostatic adenocarcinoma (PC3).
- the MTSP6 transcript was also detected in breast cancer cell line MDA-MB-231, prostate cancer cell line PC-3, but not in HT-1080 human fibrosarcoma cell line.
- MTSP6 also is expressed in mammary gland carcinoma cDNA (Clontech).
- MTSP6 also is over expressed in ovarian tumor cells.
- the MTSP7 transcript was detected in lung carcinoma (A549 cell line), leukemia (K-562 cell line) and cervical carcinoma (HeLaS3 cell line). MTSP7 is believed to be expressed in lung, colon, prostate, breast, cervical and other tumors.
- MTSP9 is, for example, expressed in esophageal tumor tissues, in lung carcinoma, in colorectal carcinoma, lymphoma, a cervical carcinoma (HeLaS3) and leukemia cell lines as well as in certain normal cells and tissues.
- MTSP9 also can be a marker for breast, prostate, cervical and colon cancer.
- MTSP9 is highly expressed in the esophagus and expressed at a low level in many other tissues.
- the MTSP9 transcript is found in kidney (adult and fetal), spleen (adult and fetal), placenta, liver (adult and fetal), thymus, peripheral blood leukocyte, lung (adult and fetal), pancreas, lymph node, bone marrow, trachea, uterus, prostate, testes, ovary and the gland organs (mammary, adrenal, thyroid, pituitary and salivary).
- MTSP9 also is expressed in esophagus tumor tissues, in a lung carcinoma and, at a lower level, in a colorectal carcinoma, lymphoma, a cervical carcinoma (HeLaS3) and leukemia cell lines.
- MTSP10 for example, is expressed in esophageal tumor tissues, in lung carcinoma, prostate cancers, pancreatic and breast cancers and in cell lines as well as in certain normal cells and tissues (see e.g., EXAMPLES for tissue-specific expression profile).
- the level of activated MTSP10 can be diagnostic of prostate, uterine, lung esophagus, or colon cancer or leukemia or other cancer.
- the expression and/or activation of MTSP10 on or in the vicinity of a cell or in a bodily fluid in a subject can be a marker for breast, prostate, lung, colon, esophageal and other cancers.
- MTSP10 transcript was detected in pancreas, lung and kidney. MTSP10 transcript was also detected in small intestine Marathon-Ready cDNA (Clontech). The MTSP10 transcript was detected in breast carcinoma (GI-101), lung carcinoma (LX-1 and GI-117), ovarian carcinoma (GI-102), and pancreatic adenocarcinoma (GI-103). The MTSP10 transcript was weakly detected in prostatic adenocarcinoma (PC3). The MTSP10 transcript was also detected in CWR22R prostate tumor grown in nude mice. No apparent signal was detected in two forms of colon adenocarcinomas (GI-112 and CX-1).
- MTSP12 transcript was detected in pancreas, lung and kidney. MTSP12 transcript was also detected in small intestine Marathon-Ready cDNA (Clontech). The MTSP12 transcript was detected in breast carcinoma (GI-101), lung carcinoma (LX-1 and GI-117), ovarian carcinoma (GI-102), and pancreatic adenocarcinoma (GI-103). The MTSP12 transcript was weakly detected in prostatic adenocarcinoma (PC3). The MTSP12 transcript was also detected in CWR22R prostate tumor grown on nude mice. No apparent signal was detected in two forms of colon adenocarcinomas (GI-112 and CX-1).
- MTSP20 is expressed in the lung, colon, cervical tumors and in leukemic cells. It may also be expressed in breast, ovarian, pancreatic, prostate and in other tumors. MTSP20 transcript was detected in liver, lymph node, cerebellum, pancreas, prostate, uterus, testis, glands (adrenal, thyroid and salivary), thymus, kidney and spleen. Lower transcript level was found in lung, placenta, bladder, ovary, digestive system, circulatory system and other parts of the the brain.
- MTSP20 is also expressed in certain tumor cell lines including lung carcinoma (A519), colorectal carcinoma (SW480), lymphoma (Raji and Daudi), cervical carcinoma (HeLaS3) and leukemia (HL-60, K-562 and MOLT-4) cell lines.
- MTSP22 is expressed in the uterine tissue, thymus, adipose tissue, and lymph node. It may also be expressed in lung, stomach, uterine, breast, ovarian, prostate and in other tumors. MTSP22 transcript was detected in some uterus tissue samples, but not in their matched tumor samples. In one of 42 uterus samples, MTSP22 is expressed in tumor and its metastatic tissues, but not in the normal tissue counterpart. MTSP22 is also expressed in some stomach tumors and lung tumors, but not in their normal tissue counterparts. MTSP22 is also expressed in the normal tissue of a pancreas matched cDNA pair. MTSP22-encoding cDNA was detected in thymus, adipose tissue, and lymph node
- MTSP25 is expressed in breast, colon, uterine, ovarian, kidney, prostate, testicular cancer tissue. It may also be expressed in lung, stomach, prostate and in other tumors. MTSP25 transcript was expressed weakly in the lymph node. In the cancer profiling array analysis, MTSP25 is highly expressed in prostate samples (in normal and cancer samples). MTSP25 was highly expressed in a kidney tumor sample, but not in its normal tissue counterpart. MTSP25 was also expressed a breast cancer samples, but not in its normal tissue counterpart. MTSP25 was expressed in normal uterus samples, but not in their tumor counterparts. MTSP25 expression was also ovarian cancer samples. Among these three samples, the expression of MTSP25 was also detected in one of the matched normal tissue counterparts. MTSP25 expression was also detected in tumor samples in colon cDNA pairs.
- PCR analysis revealed that MTSP25 cDNA was strongly detected in testis and mammary gland adenocarcinoma, weakly detected in brain, placenta, lung, spleen, prostate, small intestine, colon, and leukocyte, and very weakly detected in heart, liver and pancreas.
- Endotheliases are a class of cell surface proteases that are expressed on cells, particularly endothelial cells, particularly those proliferating endothelial cells, which are involved in a variety of proliferative processes, including undesirable angiogenesis associated with tumor growth and metastasis, and with other hyperproliferative disorders, such as restenosis, scarring, diabetic retinopathies, diseases and disorders of the anterior eye (see, U.S. application Ser. No. 09/717,473, filed Nov. 20, 2000, and International PCT application No. PCT/US00/31803).
- Endotheliases are particularly useful targets for delivery of therapeutic agents for treatment of any disorder involving aberrant angiogenesis.
- Endothelial cells play a key role in angiogenesis, which is is the generation of new blood vessels from parent microvessels.
- Angiogenesis plays a major role in the metastasis of cancer and in the pathology of a variety of other disorders.
- Controlled and uncontrolled angiogenesis proceed in a similar manner. Endothelial cells and pericytes, surrounded by a basement membrane, form capillary blood vessels. Angiogenesis begins with the erosion of the basement membrane by enzymes released by endothelial cells and leukocytes. The endothelial cells, which line the lumen of blood vessels, then protrude through the basement membrane. Angiogenic stimulants induce the endothelial cells to migrate through the eroded basement membrane. The migrating cells form a “sprout” off the parent blood vessel, where the endothelial cells undergo mitosis and proliferate. The endothelial sprouts merge with each other to form capillary loops, creating the new blood vessel.
- Angiogenesis is highly regulated by a system of angiogenic stimulators and inhibitors.
- angiogenesis stimulators include certain growth factors, cytokines, proteins, peptides, carbohydrates and lipids (Norrby (1997) APMIS 105:417-437); Polverini (1995) Crit. Rev. Oral. Biol. Med . 6:230-247).
- endogenous and exogenous angiogenesis inhibitors are known in the art (Jackson et al. (1997) FASEB 11:457-465; Norrby (1997) APMIS 105:417-437); and O'Reilly (1997) Investigational New Drugs , 15:5-13).
- Angiogenesis is essential for normal placental, embryonic, fetal and post-natal development and growth, but almost never occurs physiologically in adulthood except in very specific restricted situations. For example, angiogenesis is normally observed in wound healing, fetal and embryonal development and formation of the corpus luteum, endometrium and placenta. Angiogenesis in the adult is often associated with disease states.
- angiogenesis is involved in the manifestation or progress of various diseases, for example, various inflammatory diseases, such as rheumatoid arthritis, psoriasis, diabetic retinopathies, certain ocular disorders, including recurrence of pterygii, scarring excimer laser surgery and glaucoma filtering surgery, various disorders of the anterior eye, cardiovascular disorders, chronic inflammatory diseases, wound repair, circulatory disorders, crest syndromes, dermatological disorders (see, e.g., U.S. Pat. Nos. 5,593,990, 5,629,327 and 5,712,291) and notably cancer, including solid neoplasms and vascular tumors.
- various inflammatory diseases such as rheumatoid arthritis, psoriasis, diabetic retinopathies, certain ocular disorders, including recurrence of pterygii, scarring excimer laser surgery and glaucoma filtering surgery, various disorders of the anterior eye, cardiovascular disorders,
- Angiogenesis is essential for the growth and persistence of solid tumors and their metastases. Repressing, eliminating or modulating this activity, should impact the etiology of these diseases and serve as a point of therapeutic intervention. In the disease state, prevention of angiogenesis could avert the damage caused by the invasion of the new microvascular system. Therapies directed at control of the angiogenic processes could lead to the abrogation or mitigation of these diseases. Hence there is a need to develop therapeutics that target angiogenesis and modulate, particularly, inhibit aberrant or uncontrolled angiogenesis.
- conjugates that contain endotheliase substrates can be used to deliver therapeutic agents for the treatment of diseases including, but are not limited to, rheumatoid arthritis, psoriasis, diabetic retinopathies, other ocular disorders, including recurrence of pterygii, scarring from excimer laser surgery and glaucoma filtering surgery, various disorders of the anterior eye, cardiovascular disorders, autoimmune diseases, chronic inflammatory diseases, wounds, circulatory disorders, crest syndromes, restenosis, psoriasis and other dermatological disorders (see, e.g., U.S. Pat. Nos. 5,593,990, 5,629,327 and 5,712,291) and notably cancer, including solid neoplasms and vascular tumors.
- diseases including, but are not limited to, rheumatoid arthritis, psoriasis, diabetic retinopathies, other ocular disorders, including recurrence of pterygi
- endotheliases are two different endotheliases and variant forms thereof designated endotheliase 1 and endotheliase 2 (see SEQ ID Nos. 21-27.
- Other members of the family can be identified by probing for genes or searching libraries for genes that have sequence identity, particularly at least 40%, 60%, 80%, 90%, 95%, 98% or greater sequence identity to the protease domain of an endotheliase identified herein, or that hybridize under conditions of high stringency to the full-length of the nucleic acid encoding a protease domain of an endotheliase provided herein, and that are expressed on endothelial cells.
- Exemplary of the endotheliase are endotheliase 1 and endotheliase 2. These are expressed on endothelial cells.
- Exemplary of a full-length endotheliase 1 is one that includes the sequence of amino acids set forth in SEQ ID No. 42 (see, International PCT application No. WO 00/5006, which describes a gene it designates DESC1 that is expressed in squamous cell carcinomas and prostate tumors).
- endotheliases are expressed on endothelial cells.
- a protease domain thereof is set forth in SEQ ID NO: 22.
- RNA blot composed of 76 different human tissues (catalog number 7775-1; human multiple tissue expression (MTE) array; CLONTECH, Palo Alto, Calif.). Significant expression was observed in the esophagus, with minor expression levels in the stomach, salivary gland, pancreas, prostate, bladder, trachea and uterus.
- Northern analysis using RNA blots confirmed that the expression was restricted to the esophagus. Two transcripts (approximately 1.7 and 2 kb) were detected in the esophagus. Endotheliase 1 also is expressed in umbilical vein endothelial cells, PC3 and LnCAP cells.
- endotheliase 2-S and endotheliase 2-L Two splice variant forms of endotheliase 2 designated endotheliase 2-S and endotheliase 2-L are exemplified herein (see SEQ ID Nos. 23-26).
- the open reading frame of the nucleic acid encoding endotheliase 2-S (SEQ ID No. 23) is composed of 1,689 bp, which translates to a 562-amino acid protein (SEQ ID No. 24), while the ORF of endotheliase 2-L is composed of 2,067 bp (SEQ ID No. 25), which translates to a 688-amino acid protein (SEQ ID No. 26).
- the nucleic acid encoding the protease domain of endotheliase 2-S is composed of 729 bp which translates to a 242-amino acid protein (amino acids 321-562 of SEQ ID Nos. 23 and 24), while that of endotheliase 2-L is composed of 1,107 bp, which translates to a 368-amino acid protein (amino acids 321-688 of SEQ ID Nos. 25 and 26).
- any and all of the above-noted endotheliases and/or protease domains thereof such as those that include the sequences of amino acids in SEQ ID Nos. 22, 24, 26 and 27 or are encoded by nucleic acid that hybridize thereto under the conditions as described above are contemplated for use in the methods herein.
- proteins that include amino acid sequence changes such as those set forth in Table 1 above, and retain protease activity.
- Conjugates that are substrates for proteases on the surfaces of cells, particularly serine proteases, including type II membrane-bound serine proteases, and endotheliases are provided. Any cell surface protease, including cell-associated or localized proteases, is contemplated herein. Generally proteases expressed at high levels in active forms in essential tissues are not ideal target candidates. The proteases include those that are expressed on relatively limited numbers of cells or that are expressed at high levels in cells, such as tumor cells and endothelial cells and immune cells, that are involved in disease states or are present in diseases states in the locale of cells involved in the disease states.
- endothelial cells by virtue of their role in angiogenesis are involved in numerous proliferative disorders; immune cells are involved in many disease processes including cancers and diseases and inflammatory disorders.
- Other cell surface proteases are expressed at higher levels in certain tumors than in normal cells. Whether or not such proteases have a role in the disorder their higher expression in cells involved in a disease state is sufficient for use for targeting therapeutic agents in the conjugates provided herein.
- the conjugates which contain a therapeutic agent, such as a cytotoxic agent, is activated upon cleavage by a cell surface protease, including cell-associated and cell-localized proteses.
- a cell surface protease including cell-associated and cell-localized proteses.
- proteases are the MTSPs, such as, but not limited to, MTSP1, MTSP3, MTsP4, MTSP6, MTSP7, MTSP9, MTSP10, MTSP12, MTSP20, MTSP22, MTSP25, urokinases and endotheliases.
- the conjugates targeted to such proteases are prodrugs in that the therapeutic agent is inactive as administered and is ultimately activated in the vicinity of the targeted cell or tissue.
- cell surface proteases such as transmembrane proteases, are the intended targets, any released, shed or soluble forms of the proteases and others also can be targeted.
- the conjugates which contain a therapeutic agent, such as a cytotoxic agent, are substantially inactive prior to action by a cell surface protease, a peptidic moiety that is a substrate for a targeted cell surface protease (i.e., a peptidic substrate), and, optionally, a linker.
- the therapeutic agents in the conjugates are activated upon cleavage of the peptidic substrate of the conjugate by a cell surface protease.
- the therapeutic agents, such as cytotoxic agents are released as the free yagent, or, alternatively, are released coupled to the portion of the peptidic substrate (P1-P2-P3-etc.
- cytotoxic agents in these forms, are released in the vicinity of cells that express the proteases. Activation is effected, in certain embodiments, because the therapeutic agent, such as cytotoxic agent, following action of the cell surface protease, can cross the cell membrane or otherwise interact with the cell or tissue and exhibit therapeutic activity. In other embodiments, any remaining peptidic moieties or amino acids can be cleaved from therapeutic agent to render it active.
- the conjugates act as prodrugs because the therapeutic agents when conjugated are substantially inactive. Upon cleavage by the targeted protease, the therapeutic agent is released either in active form or in a form that is activated by the targeted cell, tissue or surrounding environment.
- the targeted agent is a cytotoxic agent and the conjugates for use in the methods and compositions provided herein have the formula:
- peptide l is a peptidic substrate for a cell surface protease or a released, shed or otherwise unbound membrane protease, such as an MTSP; s is greater than or equal to 1, or is 1 to 6, or is 1 or 2, or is 1; linker is any linker; q is greater than or equal to 0, or is 0 to 4, or is 0 or 1; the cytotoxic agent is an anti-tumor, anti-cancer or anti mitotic agent, including anti-antiangiogenic agents; and t is 1 or more, or is 1 or 2.
- the cytotoxic agent is covalently attached, optionally via a linker, to either the C-terminus or the N-terminus of the peptidic substrate.
- the therapeutic agent such as a cytotoxic agent
- the N-terminus optionally is capped.
- N-Terminal caps for use herein include, but are not limited to, acyl, sulfonyl and carbamoyl groups.
- the C-terminus is a carboxamide derivative.
- peptide l is a peptidic substrate for a cell surface protease or a soluble MTSP whereby, upon action of the protease, the conjugate, which is substantially inactive, is cleaved at the P1-P1′ bond to release a compound of the formula:
- peptide a is a truncated version of peptide l resulting from cleavage at the P1-P1′ bond.
- the conjugates for use in the methods and compositions provided herein possess two therapeutic agents, such as cytotoxic agents, which are the same or different, linked to the C-terminus and the N-terminus, respectively, optionally via linkers linker 1 and linker 2 , of a peptidic substrate for cell surface protease or a soluble MTSP.
- the conjugates have the formula:
- peptide l is a peptidic substrate for a cell surface protease, or a soluble MTSP; s is greater than or equal to 1, or is 1 to 6, or is 1 or 2, or is 1; linker 1 and linker 2 are each independently any linker and are the same or different; q and w are each independently greater than or equal to 0, or are 0 to 4, or are 0 or 1; the therapeutic agents, which are the same or different, are anti-tumor, anti-cancer or anti mitotic agents; and t and x are each independently 1 or more, or are 1 or 2.
- peptide l is a peptidic substrate for a cell surface protease or a soluble MTSP whereby, upon action of the protease, the conjugate, which is substantially inactive, is cleaved at a point on the peptidic chain to release two compounds of the formulae:
- peptide a1 and peptide a2 are N-terminal and C-terminal truncated portions, respectively, of peptide l resulting from cleavage at the P1-P1′ bond.
- conjugates for use in the compositions and methods provided herein have formula I:
- l is 0 or 1; when l is 0, j, i, p and m are 0; when l is 1, j is 0 or 1; when j is 0, i, p and m are 0; when j is 1, i is 0 or 1; when i is 0, p and m are 0; when i is 1, p is 0 or 1; when p is 0, m is 0; when p is 1, m is 0 or 1;
- u, k and r are selected as follows:
- u is 0 or 1; when u is 0, k and r are 0; when u is 1, k is 0 or 1; when k is 0, r is 0; when k is 1, r is 0 or 1;
- n is 0 or 1;
- X n is hydrogen, or an acyl, sulfonyl or carbamoyl cap; and P6 to P3′ are amino acid residues, as defined below.
- the P6 to P3′ residues are linked by peptide bonds or peptide bond surrogates.
- the P6 to P3′ portion of the conjugate is a peptidic substrate, as defined herein.
- conjugates for use in the compositions and methods provided herein have formula II:
- l is 0 or 1; when l is 0, j, i, p and m are 0; when l is 1, j is 0 or 1; when j is 0, i, p and m are 0; when j is 1, i is 0 or 1; when i is 0, p and m are 0; when i is 1, p is 0 or 1; when p is 0, m is 0; when p is 1, m is 0 or 1;
- u, k and r are selected as follows:
- u is 0 or 1; when u is 0, k and r are 0; when u is 1, k is 0 or 1; when k is 0, r is 0; when k is 1, r is 0 or 1;
- n is 0 or 1;
- X c together with the carbonyl group of the amino acid residue to which it is attached, forms a carboxylic acid or a carboxamide group; and
- P6 to P3′ are amino acid residues, as defined below.
- the P6 to P3′ residues are linked by peptide bonds or peptide bond surrogates.
- the P6 to P3′ portion of the conjugate is a peptidic substrate, as defined herein.
- conjugates for use in the compositions and methods provided herein have formula III:
- Z 1 and Z 2 are each therapeutic agents and are the same or different;
- L 1 and L 2 are each linkers and are the same or different;
- l, j, i, p and m are selected as follows:
- l is 0 or 1; when l is 0, j, i, p and m are 0; when l is 1, j is 0 or 1; when j is 0, i, p and m are 0; when j is 1, i is 0 or 1; when i is 0, p and m are 0; when i is 1, p is 0 or 1; when p is 0, m is 0; when p is 1, m is 0 or 1;
- u, k and r are selected as follows:
- u is 0 or 1; when u is 0, k and r are 0; when u is 1, k is 0 or 1; when k is 0, r is 0; when k is 1, r is 0 or 1;
- n and v are each independently 0 or 1; and P6 to P3′ are amino acid residues, as defined below.
- the P6 to P3′ residues are linked by peptide bonds or peptide bond surrogates.
- the P6 to P3′ portion of the conjugate is a peptidic substrate, as defined herein.
- conjugates for use in the compositions and methods provided herein have formula IV:
- l is 0 or 1; when l is 0, j, i, p and m are 0; when l is 1, j is 0 or 1; when j is 0, i, p and m are 0; when j is 1, i is 0 or 1; when i is 0, p and m are 0; when i is 1, p is 0 or 1; when p is 0, m is 0; when p is 1, m is 0 or 1;
- u, k, r and s are selected as follows:
- u is 0 or 1; when u is 0, k, r and s are 0; when u is 1, k is 0 or 1; when k is 0, r and s are 0; when k is 1, r is 0 or 1; when r is 0, s is 0; when r is 1, s is 0 or 1;
- n is 0 or 1;
- X n is hydrogen, or an acyl, sulfonyl or carbamoyl cap; and P6 to P4′ are amino acid residues, as defined below.
- the P6 to P4′ residues are linked by peptide bonds or peptide bond surrogates.
- the P6 to P4′ portion of the conjugate is a peptidic substrate, as defined herein.
- the conjugates for use in the compositions and methods provided herein have formula V:
- l is 0 or 1; when l is 0, j, i, p and m are 0; when l is 1, j is 0 or 1; when j is 0, i, p and m are 0; when j is 1, i is 0 or 1; when i is 0, p and m are 0; when i is 1, p is 0 or 1; when p is 0, m is 0; when p is 1, m is 0 or 1;
- u, k, r and s are selected as follows:
- u is 0 or 1; when u is 0, k, r and s are 0; when u is 1, k is 0 or 1; when k is 0, r and s are 0; when k is 1, r is 0 or 1; when r is 0, s is 0; when r is 1, s is 0 or 1;
- n is 0 or 1;
- X c together with the carbonyl group of the amino acid residue to which it is attached, forms a carboxylic acid or a carboxamide group; and
- P6 to P4′ are amino acid residues, as defined below.
- the P6 to P4′ residues are linked by peptide bonds or peptide bond surrogates.
- the P6 to P4′ portion of the conjugate is a peptidic substrate, as defined herein.
- conjugates for use in the compositions and methods provided herein have formula VI:
- Z 1 and Z 2 are each therapeutic agents and are the same or different;
- L 1 and L 2 are each linkers and are the same or different;
- l, j, i, p and m are selected as follows:
- l is 0 or 1; when l is 0, j, i, p and m are 0; when l is 1, j is 0 or 1; when j is 0, i, p and m are 0; when j is 1, i is 0 or 1; when i is 0, p and m are 0; when i is 1, p is 0 or 1; when p is 0, m is 0; when p is 1, m is 0 or 1;
- u, k, r and s are selected as follows:
- u is 0 or 1; when u is 0, k, r and s are 0; when u is 1, k is 0 or 1; when k is 0, r and s are 0; when k is 1, r is 0 or 1; when r is 0, s is 0; when r is 1, s is 0 or 1;
- n and v are each independently 0 or 1; and P6 to P4′ are amino acid residues, as defined below.
- the P6 to P4′ residues are linked by peptide bonds or peptide bond surrogates.
- the P6 to P4′ portion of the conjugate is a peptidic substrate, as defined herein.
- the peptidic substrates contemplated for use in the conjugates are substrates for the targeted cell surface protease or a soluble, shed or released form thereof, and contain a sufficient number of amino acid residues to render any therapeutic agent in the conjugate substantially inactive.
- the therapeutic agent is, for example, doxorubicin
- the conjugate is substantially inactive by virtue of the inability of the conjugated therapeutic agent to cross the cell membrane.
- the peptidic substrate contains at least 1, 2, 3, 4 or 5 amino acid residues, and can contain up to nine or ten residues. Longer peptidic substrates can be used in the conjugates as long as upon cleavage, the resulting therapeutic agent or therapeutic agent-amino acid or -peptidic moiety conjugate exhibits the desired therapeutic effect in vivo and in vitro.
- exemplary peptidic substrates for use in the conjugates provided herein possess at least one amino acid (P1), two amino acids (P1-P1′), three amino acids (P2-P1-P1′) and typically contain four, five or six amino acid residues (P3-P2-P1-P1′, P4-P3-P2-P1-P1′ or P4-P3-P2-P1-P1′-P2′), where the P1-P1′ bond is the site of cleavage of cell surface protease, or a soluble, shed or released form thereof, including, but not limited to, a cell surface protease, such as a serine protease, including, for example, but not limited to, uPA bound to its receptor, MTSPs and endotheliases.
- P1-P1′ two amino acids
- P2-P1-P1′ typically contain four, five or six amino acid residues
- the peptidic substrates optionally further possess a P5, P6 or P3′ amino acid residue, and, in certain embodiments, possess P7, P8, P9, P10, P4′, P5′, P6′ residues.
- the peptidic substrates for use in the conjugates provided herein are penta-, hexa-, hepta-, octa- and nona-peptidic substrates, and can contain 10, 11, 12, 13, 14, 15 or more residues as long as, upon cleavage of the conjugate by the protease, the resulting therapeutic agent or therapeutic agent-amino acid or -peptidic moiety conjugate exhibits the desired therapeutic effect in vivo and in vitro.
- the peptidic substrates are conjugated to the therapeutic agent (or to a linker to which the therapeutic agent is linked) via the C-terminal residue (i.e., P1′, P2′ or P3′), or the N-terminal residue (i.e., P6, P5 or P4), or optionally an internal residue.
- the peptidic substrates for example, can be straight chains, but can be cyclized or include cyclized portions.
- the peptidic substrate optionally possesses a cap, such as an acyl or carbamoyl cap at the N-terminus.
- the peptidic substrate further possess a terminal group, such as a carboxamide group, at the C-terminus.
- the conjugates can contain a plurality of peptidic substrates and a plurality of therapeutic agents.
- conjugation to the therapeutic agent(s) or linker linked thereto can be via the C-terminal and N-terminal residues of the peptidic substrate.
- substrates can be designed based upon known specificities of other proteases. For example, the specificities of trypsin-like and trypsin family members can aid in design of possible substrates. The following summarizes substrate preferences for particular serine proteases (see, e.g., Harris et al. (2000) PNAS 97(14):7754-7759).
- Typical protocols for preparation of the conjugates can include the steps of: 1) identification of a targeted protease; 2) expression and assay development; 3) substrate selection, such as, for example, by testing chromogenic or fluorogenic substrates to identify those cleaved by a selected target protease, by use of substrate phage display to identify peptidic substrates cleaved by a targeted protease, by use of a natural protein or peptide substrate or a natural inhibitor of the protease, and by use of combinatorial libraries to identify substrates cleaved by a targeted protease; 4) synthesis of conjugates containing the identified substrate; and 5) biological evaluation thereof, including, but not limited to, in vitro assays, cell culture assays, biological assays, and in vivo animal models (see, e.g., EXAMPLE 10).
- a conjugate can be designed by any methods known to those of skill in the art. The following provides an exemplary protocol. First, a series of commercially available chromogenic and fluorogenic peptidic substrates can be tested for cleavage by the protease of interest (see Examples for lists of exemplary chromogenic and fluorogenic substrates and the table below). The peptidic portion of these substrates occupies the unprimed binding sites of the protease while the reporter group is located on the primed side of the scissle bond. Effective conjugates can then be designed based on the structure of the substrates that are efficiently cleaved by the protease.
- the peptidic portion of these efficiently cleaved substrates can be used as the unprimed region of the conjugate, and Ser-therapeutic agent, such as a cytotoxic agent (e.g., doxorubicin), Ser-Leu-therapeutic agent or Ser-Ser-Leu-therapeutic agent can be used as the primed region of the conjugate. Cleavage of these conjugate prodrugs releases either Ser-therapeutic agent, Ser-Leu-therapeutic agent or Ser-Ser-Leu-therapeutic agent compounds.
- the Ser in the released Ser-therapeutic agent may be replaced by other amino acid residues including, but not limited to, Ala, hSer, Abu, Thr, Met, nLeu and Val.
- the amino acid residue conjugated to the therapeutic agent possesses a hydrophobic side chain.
- Such amino acid residues include, but are not limited to, Leu, Abu, nLeu, nVal, CHA, hCHA, (hex)Gly, (allyl)Gly, (propargyl)Gly and (cyclopropyl)Ala.
- the amino acid residue conjugated to the therapeutic agent possesses a side chain that is not sterically bulky.
- Such amino acid residues include, but are not limited to, Gly and Ala.
- conjugate prodrug for a protease substrate is to use substrate phage display to elucidate optimal subsite occupancy for the protease. The resulting information can then be used to design the peptidic, unprimed portion of the conjugate.
- the primed region of the conjugate can be fixed as Ser-therapeutic agent, Ser-Leu-therapeutic agent or Ser-Ser-Leu-therapeutic agent.
- a third approach to design an effective prodrug conjugate involves the use of combinatorial fluorogenic substrate libraries to determine optimal residues for the unprimed region of a protease substrate. These selected sequences can then be used as the unprimed portion of the conjugate prodrug and, and Ser-therapeutic agent, (e.g., doxorubicin), Ser-Leu-therapeutic agent or Ser-Ser-Leu-therapeutic agent can be used as the primed region of the conjugate.
- Ser-therapeutic agent e.g., doxorubicin
- Ser-Leu-therapeutic agent e.g., Ser-Leu-therapeutic agent
- Ser-Ser-Leu-therapeutic agent can be used as the primed region of the conjugate.
- sequences including GSGR were based on or dervied from substrate phage display experiments using u-PA as the taret protease.
- seqeuence sequences in natural substrates or natural inhibitors of a protease target, such as uPA, including VSAR, PGR (from P3-P1 of plasminogen) and related sequences were used in design of u-PA-targetd conjugates.
- sequences from chromgenic substrates, such as D-HHT-Gly-Arg, and related sequences were used for design of ET-1-targeted conjugates.
- Chromogenic/Fluorogenic Substrates Chromogenic/fluorogenic substrates Enzyme Substrate Structure MTSP1 Spectrozyme t-PA CH 3 SO 2 -D-HHT-Gly-Arg-pNA.AcOH MTSP1 S 2765 N- ⁇ -Z-D-Arg-Gly-Arg-pNA.2HCl MTSP3 Spectrozyme fXIIa H-D-CHT-Gly-Arg-pNA.2AcOH MTSP4 a Spec PL H-D-Nle-HHT-Lys-pNA.2AcOH MTSP5 S 2765 N- ⁇ -Z-D-Arg-Gly-Arg-pNA.2HCl MTSP6 spectrozyme t-PA CH 3 SO 2 -D-HHT-Gly-Arg-pNA.AcOH MTSP7 S 2366 pyroGlu-Pro-Arg-pNA.HCl MTSP9 Pefachrome
- a protease for a coupled assay, the ability of the protease to activate an enzyme, such as plasminogen or trypsinogen is tested.
- a protease is incubated with a zymogen, such as plasminogen or trypsinogen, in the presence of a labelled known substrate, such as lys-plasminogen or Spec PL (for plasmin), for the zymogen. If protease activates the zymogen, the activated enzyme, such as plasmin and trypsin, will degrade the substrate, thereby changing the spectral properties of the substrate.
- a zymogen such as plasminogen or trypsinogen
- peptidic substrates for cleavage by proteases such as MTSP1 (or matriptase), endotheliase 1 and urokinase, and a general discussion of properties of the residues.
- proteases such as MTSP1 (or matriptase), endotheliase 1 and urokinase
- peptidic substrates for cleavage by other cell surface proteases, or a soluble, shed or released form thereof can be similarly designed by identifying peptidic substrates for the selected protease and then preparing conjugates that contain such peptidic substrates.
- Amino acid residues for use at the P1 position of the peptidic substrates for use in the conjugates provided herein include Arg, Arg surrogates and Lys.
- Arg surrogates include unnatural amino acids that possess a group or moiety that functions in substantially the same way as the naturally occurring side chain of arginine to achieve substantially the same result (e.g., acting as the P1 residue in a substrate for a MTSP1, urokinase or endotheliase).
- Arg surrogates include, but are not limited to, ⁇ -amino acids that possess as the side chain any of the following: the side chain of homoarginine; guanidinoaminopropyl; guanidinoaminoethyl; (Me) 2 arginine side chain; (Et) 2 arginine side chain; (4-aminomethyl)phenylmethyl; 4-amidinophenylmethyl; 4-guanidinophenylmethyl; or the Arg surrogate is a conformationally constrained arginine analog such as:
- z is 0 or 1 (see, e.g., Webb et al. (1991) J. Org. Chem . 56:3009); or the side chain is a conformationally constrained arginine side chain analog such as:
- d is an integer from 0 to 5, or 1 to 3; and W is N or CH; or a mono- or di-substituted N-alkyl derivative of the above groups, where alkyl is, in certain embodiments, lower alkyl, such as, for example, methyl.
- the P1 residue is Arg.
- the P2 residue is selected from Phe, Ser, Gly, Ala, Ser(OMe), hSer, 1-methylHis, 3-methylHis, His, nVal, nLeu, Abu, (hS)Gly, Thr, Aib, CHA and Tyr.
- the P2 residue is selected from Phe, Ser, Gly and Ala.
- the P2 residue is Ser or Ala.
- the P2 residue is Gly or Ala.
- Amino acid residues for use at the P3 position of the conjugates provided herein include Arg, Lys, Gln, Quat, Arg surrogates, Ser, Thr, hSer, dSer, Pro, (hS)Gly, Tyr, 4,4-dimethylThr, Asn, Met(O 2 ), Quat 2 , Quat 3 , Quat 4 and Quat 5 .
- the P3 residue is selected from Arg, Lys, Gln, Quat and Arg surrogates.
- Arg surrogates include those described above for the P1 residue.
- the P3 residue is Gin or Ser.
- the P4 residue is selected from Pro, Arg, Ser, Ala, Lys, Gly, nLeu, Leu, Tyr, Glu, Phe, Val, N,N-dimethylGly, ⁇ -Ala, Cys(Me), Gin, t-butylGly and nVal.
- the P4 residue is selected from Pro, Arg, Ser, Ala, Lys, Gly, nLeu, Leu, Tyr, Glu, Phe and Val.
- the P4 residue is selected from Pro, Arg, Ser, Ala, Lys, Gly, nLeu, Phe or Val.
- the P4 residue is Arg or Gly.
- the peptidic substrates used in the conjugates contain a P5 and, optionally, a P6 residue.
- P5 residues include Ile, Arg and Arg surrogates.
- P5 residues include Arg and Arg surrogates.
- Arg surrogates include those described above for the P1 residue.
- P6 residues include, for example, Leu, Val and Arg.
- P6 residues include, for example, Leu.
- the P1′ residue of the conjugates provided herein is Gly, Ser, Ala, Leu, Ile, d-Ile, nLeu, Val, nVal, Aib, Abu, Met, 6-aminohexanoyl, Thr or hSer.
- the P1′ residue of the conjugates provided herein is Gly, Ser, Ala, Leu, Ile, d-Ile, nLeu, Val, nVal, Aib, Abu, Met or 6-aminohexanoyl.
- the P1′ residue is Ser, Ala, hSer, Abu, Thr, Met, nLeu or Val.
- the P1′ residue is Gly or Ala.
- the P1′ residue is Ser, Ala or Gly. In another embodiment, the P1′ residue is Leu, Abu, nLeu, nVal, CHA, hCHA, (hex)Gly, (allyl)Gly, (propargyl)Gly or (cyclopropyl)Ala. In certain embodiments herein, the P1′ residue is Ala, Ser, Gly, Ile or d-Ile.
- the conjugates provided herein possess a P2′ residue.
- P2′ residues for use herein include, but are not limited to, Gly, Ser, Ala, Leu, Ile, d-Ile, nLeu, Val, nVal, Aib, Abu, Met, 6-aminohexanoyl, hCHA, CHA, hexylGly, allylGly and Phe.
- P2′ residues for use herein include, but are not limited to, Gly, Ser, Ala, Leu, Ile, d-Ile, nLeu, Val, nVal, Aib, Abu, Met and 6-aminohexanoyl.
- the P2′ residue is Ser, hSer, Abu, nLeu, nVal, CHA, hCHA, (allyl)Gly or (hexyl)Gly.
- the P2′ residue is Gly or Ala.
- the P2′ residue is Leu, Abu, nLeu, nVal, CHA, hCHA, (hex)Gly, (allyl)Gly, (propargyl)Gly or (cyclopropyl)Ala.
- the P2′ residues are Ala, Gly, Ile or d-Ile.
- the peptidic substrates used in the conjugates provided herein include a P3′ residue.
- P3′ residues for use herein include, but are not limited to, Gly, Ser, Ala, Leu, Ile, nLeu, Val, nVal, Aib, Abu, Met, 6-aminohexanoyl, CHA and allylGly.
- the P2′ residue is Ser, hSer, Abu, nLeu, nVal, CHA, hCHA, (allyl)Gly or (hexyl)Gly.
- P3′ residues for use herein include, but are not limited to, Gly, Ser, Ala, Leu, Ile, nLeu, Val, nVal, Aib, Abu, Met and 6-aminohexanoyl.
- the P3′ residue is Gly or Ala.
- the P3′ residue is Leu, Abu, nLeu, nVal, CHA, hCHA, (hex)Gly, (allyl)Gly, (propargyl)Gly or (cyclopropyl)Ala.
- the peptidic substrates used in the conjugates provided herein include a P4′ residue.
- P4′ residues for use herein include, but are not limited to, Gly, Ser, Ala, Leu, Ile, nLeu, Val, nVal, Aib, Abu, Met, 6-aminohexanoyl, CHA and allylGly.
- P4′ residues for use herein include, but are not limited to, Gly, Ser, Ala, Leu, Ile, nLeu, Val, nVal, Aib, Abu, Met and 6-aminohexanoyl.
- the P4′ residue is Gly or Ala.
- the P4′ residue is Leu, Abu, nLeu, nVal, CHA, hCHA, (hex)Gly, (allyl)Gly, (propargyl)Gly or (cyclopropyl)Ala. In another embodiment, the P4′ residue is Leu.
- the N-terminus of the peptidic substrate optionally is capped with an acyl, sulfonyl or carbamoyl derivative.
- the cap is chosen, in certain embodiments, to increase the hydrophilicity of the conjugate.
- a non-hydrophilic N-terminal cap such as an acetyl group, can be used.
- the N-terminal amino acid is modified with a hydrophilic blocking group.
- blocking groups are chosen based upon the presence of hydrophilic functionality. Such blocking of the terminal amino group can also reduce or eliminate the enzymatic degradation of such peptidyl therapeutic agents by the action of exogenous amino peptidases which are present in the blood plasma of warm blooded animals.
- N-Terminal blocking groups that increase the hydrophilicity of the conjugates and therefore increase the aqueous solubility of the conjugates include, but are not limited to, hydroxylated alkanoyl, polyhydroxylated alkanoyl, polyethylene glycol, glycosylates, sugars and crown ethers.
- the N-terminal blocking group is one of the following:
- R 1 and R 2 are selected from (i) or (ii) as follows:
- R 1 and R 2 are each independently:
- substituted C 1 -C 6 alkyl wherein the substituent on the substituted C 1 -C 6 alkyl is selected from unsubstituted or substituted aryl, unsubstituted or substituted heterocyclyl, C 3 -C 10 cycloalkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, R 3 O—, R 4 S(O) e NH—, R 3 C(O)NR 3 —, (R 3 ) 2 NC(O)—, (R 3 ) 2 N—C(NR 3 )—, —CN, R 3 C(O)—, —N 3 , —N(R 3 ) 2 , and R 4 OC(O)—NR 3 —; or
- R 3 is selected from: hydrogen, unsubstituted or substituted aryl, unsubstituted or substituted heterocyclyl, C 1 -C 6 alkyl and C 3 -C 10 cycloalkyl;
- R 4 is selected from: unsubstituted or substituted aryl, unsubstituted or substituted heterocyclyl, C 1 -C 6 alkyl and C 3 -C 10 cycloalkyl;
- a is 1, 2, 3 or 4;
- c is 0 to 10, provided that if b is zero, c is 1 to 10;
- f is 3, 4 or 5.
- R 1 and R 2 are each independently hydrogen, OH, C 1 -C 6 alkyl, C 1 -C 6 alkoxy, C 1 -C 6 aralkyl or aryl.
- a is 1, 2, 3 or 4;
- b is 0 or an integer between 1 and 100; and
- c is 0 to 10, provided that if b is 0, c is 1 to 10.
- the N-terminal cap (X n ) is hydrogen, or (i), (ii), (iii) or (iv) as follows:
- X n is R 30 O—C(O)—, R 31 R 32 N—C(O)—, R 33 (CH 2 ) k C(O)— or H—C(O)—; where k is an integer from 1 to 4, or is 1 or 2;
- R 30 is alkyl, aryl, heteroaryl, aralkyl or heteroaralkyl;
- R 31 and R 32 are each independently hydrogen, alkyl, aryl, heteroaryl, aralkyl, or heteroaralkyl;
- R 33 is hydrogen, hydroxy, alkyl, alkenyl, alkynyl, alkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, aralkyl, aralkoxy, heteroaralkyl or heteroaralkoxy.
- X n is hydrogen, acetyl, hydroxyacetyl, 2,3-dihydroxypropionyl, 2,3,4-trihydroxybutanoyl, PEG(1), PEG(2), PEG(4), PEG(6), PEG(14), PEG(15), PEG(16), PEG(17), PEG(18) or PEG(19).
- X n is hydrogen, acetyl, hydroxyacetyl, succinyl, quinyl, gallyl, 4-imidazolylacetyl, cotininyl, 3-phosphonylpropionyl, gulonyl, 4-phosphonylbutyryl, glutaryl, ethoxysquaryl or PEG(2).
- X n is hydrogen, acetyl, —C(O)NH 2 , HOCH 2 CH 2 C(O)—, diaminopropanoyl, or NH 2 —(CH 2 ) 5 —C(O)—.
- X n is hydrogen, acetyl, succinyl, glutaryl, PEG(2) or malonyl.
- X n is hydrogen, acetyl, succinyl, glutaryl, PEG(2), malonyl, methoxycarbonyl, phenylsulfonyl, 3-methoxypropanoyl, ethoxycarbonyl, isobutoxycarbonyl, benzyloxycarbonyl, tert-butoxycarbonyl, 4-oxopentanoyl, 2-(2-methoxyethoxy)ethoxy)acetyl, 3,4-methylenedioxyphenylacetyl, 2-pyridylacetyl, phenoxyacetyl, phenylacetyl, methoxyacetyl, 2-methoxyethoxycarbonyl, 2-methoxyethoxyacetyl, 3-phenyl-2-hydroxypropanoyl, pent-4-ynoyl
- X c together with the carbonyl group to which it is attached, forms a carboxamide derivative of formula —C(O)NR d R e , where R d and R e are selected from (i) or (ii) as follows:
- R d and R e are each independently hydrogen, C 1 -C 6 -alkyl, —C 1 -C 6 -alkyl-OH, —C 1 -C 6 -alkyl-di-OH, —C 1 -C 6 -alkyl-tri-OH and
- R d and R e are not hydrogen or C 1 -C 6 -alkyl
- c is 0 or 1, provided that if b is zero, c is 1.
- R d is hydrogen and R e is 2-hydroxyethyl.
- the conjugates optionally contain a linker (i.e., L, L 1 or L 2 of formulae I, II and III) that covalently binds the peptidic substrate to the therapeutic agent.
- the linkers are any that result in a conjugate in which the peptidic portion is a substrate for a cell surface protease and the therapeutic agent is substantially inactive when in the conjugate and is released in active form or in a form subsequently activated by the cell, tissue or environment of the targeted tissue.
- the linker can include of carbohydrate, peptide, diamine, arylamine, and/or hydrocarbon core structures.
- Linkers are desirably synthetically accessible, provide shelf-stable products, and do not possess any intrinsic biological activity that interferes with the conjugates activity. They can add desirable properties such as increasing solubility or serving to aid in trafficking the cleaved therapeutic agent in the cell.
- some linkers will be enzymatically cleaved in vitro and in vivo, and fragment to release active therapeutic agent or activatable therapeutic agent.
- the linker is, for example, a sugar and/or a peptide, such the aminosugar daunosamine.
- linkers for use herein include, but are not limited to, a biscarbonyl alkyl diradical whereby an amine moiety on the therapeutic agent is connected with the linker unit to form an amide bond and the amino terminus of the peptidic substrate is connected with the other end of the linker unit also forming an amide bond.
- a diaminoalkyl diradical linker unit whereby a carbonyl moiety on the cytotoxic agent is covalently attached to one of the amines of the linker unit while the other amine of the linker unit is covalently attached to the C-terminus of the peptidic substrate, also can be useful.
- linker units which are stable to the physiological environment when not in the presence of a cell surface protease, but are cleavable upon the cleavage of the cell surface protease proteolytic cleavage site, are intended for use herein. Furthermore, linker units can be utilized that, upon cleavage of the cell surface protease proteolytic cleavage site, remain attached to the therapeutic agent but do not significantly decrease the therapeutic activity of such a post-cleavage therapeutic agent derivative when compared with an unmodified therapeutic agent.
- the linker is a diamine containing a cyclic alkyl moiety and, in certain embodiments, the diamine contains a bicycloalkylene moiety.
- diamine linkers include, but are not limited to, 1,4-bis(aminomethyl)cyclohexane, 1,4-bis(aminomethyl)-cycloheptane, 1,3-bis(aminomethyl)cyclopentane, 1-amino-4-(aminomethyl)cyclohexane, 1,4-diaminocyclohexane and 1,4-bis(aminomethyl)-bicyclo[2.2.2]octane.
- linkers include 1, ⁇ -diaminoalkanes, including, but not limited to, 1,3-diaminopropane, and 1, ⁇ -dicarbonylalkanes, including, but not limited to, oxalic, malonic, succinic, glutaric, adipic and pivalic acids.
- linkers for use in the conjugates provided herein include self-eliminating linkers such as those of the following formulae:
- the conjugates are intended for modifying a variety of biological responses.
- the therapeutic agents are any agents, including proteins and polypeptides, small molecules and other molecules that possess or potentiate a desired biological activity.
- Such molecules include cytotoxic agents, such as, but are not limited to, a toxin such as abrin, ricin A, pseudomonas exotoxin, shiga toxin, diphtheria toxin and other such toxins and toxic portions and/or subunits or chains thereof; proteins such as, but not limited to, tumor necrosis factor, ⁇ -interferon, ⁇ -interferon, nerve growth factor, platelet derived growth factor, tissue plasminogen activator; or, biological response modifiers such as, for example, lymphokines, interleukin-I (IL-1), interleukin-2 (IL-2), interleukin-6 (IL-6), granulocyte macrophage colony stimulating factor (GMCSF), granulocyte colony stimulating factor (G-CSF), erythrop
- therapeutic agents include, but are not limited to, anti-tumor, anti-angiogenic, pro-apoptotic, anti-cancer and anti-mitotic agents. These are conjugated, optionally via a linker, to a substrate, such as peptidic substrate, which is a substrate for the protease.
- cytotoxic agents that include, in general, but are not limited to, alkylating agents, toxins, antiproliferative agents and tubulin binding agents.
- Classes of cytotoxic agents for use herein include, for example, the anthracycline family of drugs, the vinca drugs, the mitomycins, the bleomycins, the cytotoxic nucleosides, the pteridine family of drugs, diynenes, the maytansinoids, the epothilones, the taxanes and the podophyllotoxins.
- Exemplary members of those classes include, for example, doxorubicin, carminomycin, daunorubicin, aminopterin, methotrexate, methopterin, dichloro-methotrexate, mitomycin C, porfiromycin, 5-fluorouracil, 6-mercaptopurine, cytosine arabinoside, podophyllotoxin, or podophyllotoxin derivatives such as etoposide or etoposide phosphate, melphalan, vinblastine, vincristine, leurosidine, vindesine, leurosine, maytansinol, epothilone A or B, taxotere, taxol and the like.
- Such therapeutic agents include estramustine, cisplatin, combretastatin and analogs, and cyclophosphamide.
- estramustine cisplatin
- combretastatin and analogs cyclophosphamide.
- One skilled in the art can make chemical modifications to the desired therapeutic agent in order to make reactions of that compound more convenient for purposes of preparing the conjugates.
- Particular therapeutic agents include the following drugs.
- drugs One skilled in the art understands that these structural formulae are exemplary only and that such compounds or derivatives or analogs thereof have acquired in the art different generic or trivial names.
- R 12 is amino or hydroxy
- R 7 is hydrogen or methyl
- R 8 is hydrogen, fluoro, chloro, bromo or iodo
- R 9 is hydroxy or a moiety which completes a salt of the carboxylic acid.
- R 13 is hydrogen or methyl
- R 14 is methyl or thienyl or a phosphate salt thereof.
- R 15 is H, CH 3 or CHO
- R 18 is H, and one of R 16 and R 17 is ethyl and the other is H or OH;
- conjugates provided herein where the therapeutic agent is the vinca alkaloid vinblastine include those of formula:
- peptidic substrate is as described above for formulae I and II;
- L is a linker such as —NH—(CH 2 ) u —T—(CH 2 ) u —NH—;
- X n is
- R 1 and R 2 are independently hydrogen, OH, C 1 -C 6 alkyl, C 1 -C 6 alkoxy, C 1 -C 6 aralkyl and aryl;
- R 1a is C 1 -C 6 -alkyl, hydroxylated C 3 -C 8 -cycloalkyl, polyhydroxylated C 3 -C 8 -cycloalkyl, hydroxylated aryl, polyhydroxylated aryl or aryl,
- R 19 is hydrogen, (C 1 -C 3 alkyl)-CO, or chlorosubstituted (C 1 -C 3 alkyl)-CO;
- T is selected from cyclopentyl, cyclohexyl, cycloheptyl or bicyclo[2.2.2]octanyl;
- a is 1, 2, 3 or 4;
- b is zero or an integer between 1 and 100;
- c is 0 or 1, provided that if b is zero, c is 1;
- g is 1, 2 or 3;
- u is 0, 1, 2 or 3;
- R 21 is a base of one of the formulae:
- R 22 is hydrogen, methyl, bromo, fluoro, chloro or iodo
- R 23 is —OH or —NH2
- R 24 is hydrogen, bromo, chloro or lodo.
- R a is —CH 3 , —CH 2 OH, —CH 2 OCO(CH 2 ) 3 CH 3 , or —CH 2 OCOCH(OC 2 H 5 ) 2 ;
- R b is —OCH 3 , —OH or —H;
- R c is —NH 2 , —NHCOCF 3 , 4-morpholinyl, 3-cyano-4-morpholinyl, 1-piperidinyl, 4-methoxy-1-piperidinyl, benzylamine, dibenzylamine, cyanomethylamine, or 1-cyano-2-methoxyethyl amine;
- R 5 is —OH —OTHP or —H
- R 6 is —OH or —H provided that R6 is not —OH when R 5 is —OH or —OTHP.
- the therapeutic agent when the therapeutic agent is doxorubicin, it is conjugated to the peptidic substrate via the amino group of the aminoglycoside moiety of doxorubicin.
- R is PhC(O) or t-BuOC(O).
- Ribosome-inactivating proteins which include ricin, abrin and saporin, are plant proteins that catalytically inactivate eukaryotic ribosomes. RIPS inactivate ribosomes by interfering with the protein elongation step of protein synthesis.
- the RIP saporin hereinafter also referred to as SAP
- SAP RIP saporin
- conjugates for use herein include the following:
- H-D-Pro-Phe-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 87);
- H-D-Nle-HHT-Lys-Ala-Ala-(therapeutic agent) (SEQ ID NO: 94);
- H-D-CHT-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 100);
- conjugates for use in the compositions and methods provided herein include:
- conjugates for use in the compositions and methods provided herein include:
- the conjugates are Ac-Leu-Arg-Ala-Quat-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 129); Ac-Leu-Arg-Ala-Quat-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 130); Ac-Leu-Arg-Ser-Quat-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 131); and Ac-Leu-Arg-Ser-Quat-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 132).
- conjugates for use herein include the following:
- N-p-tosyl-Gly-Pro-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 147);
- H-D-Ile-Pro-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 152);
- H-D-Pro-Phe-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 154);
- H-D-Phe-Pip-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 159);
- H-D-Val-Leu-Lys-Ser-Leu-(therapeutic agent) (SEQ ID NO: 160);
- H-D-Nle-HHT-Lys-Ser-Leu-(therapeutic agent) (SEQ ID NO: 161);
- H-D-HHT-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 166);
- H-D-CHT-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 167);
- conjugates for use in the compositions and methods provided herein include:
- conjugates provided herein include:
- conjugates for use in the compositions and methods provided herein include:
- the conjugates are Ac-Leu-Arg-Ala-Quat-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 207); Ac-Leu-Arg-Ala-Quat-Ala-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 208); Ac-Leu-Arg-Ser-Quat-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 209); and Ac-Leu-Arg-Ser-Quat-Ala-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 210).
- conjugates for use herein include the following:
- N-p-tosyl-Gly-Pro-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 225); Benzoyl-Val-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 226); CH 3 SO 2 -D-HHT-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 227);
- H-D-Ile-Pro-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 230);
- H-D-Pro-Phe-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 232);
- H-D-Val-Leu-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 233);
- H-D-Val-Leu-Lys-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 238);
- H-D-Nle-HHT-Lys-Ser-Ser-Leu-(therapeutic agent) SEQ ID NO: 239
- H-D-HHT-Ala-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 244);
- conjugates provided herein include:
- conjugates provided herein are selected from:
- MeOCO-Quat2-G-R-S-L-(therapeutic agent) (SEQ ID NO: 483);
- MeOCO-Quat-G-R-S-L-(therapeutic agent) (SEQ ID NO: 485);
- MeOCO-Quat4-G-R-S-L-(therapeutic agent) (SEQ ID NO: 486);
- MeOCO-Quat5-G-R-S-L-(therapeutic agent) (SEQ ID NO: 487);
Abstract
Description
- Benefit of priority under 35 U.S.C. §119(e) to U.S. provisional application Serial No. 60/293,267, filed May 23, 2001, to Edwin L. Madison, Joseph Edward Semple and George P. Vlasuk, entitled “CONJUGATES ACTIVATED BY CELL SURFACE PROTEASES AND THERAPEUTIC USES THEREOF” is claimed. The subject matter of the above-referenced application is incorporated by reference in its entirety.
- Conjugates, compositions and methods for localized delivery of therapeutic agents for treating a variety of disorders, such as, proliferative diseases, autoimmune diseases, infectious diseases and inflammatory diseases, are provided. The conjugates, which act as prodrugs, contain therapeutic agents and peptidic substrates that are cleaved by cell surface proteases to release therapeutic agents in the vicinity of the targeted cells.
- Effective treatment of cancer and other proliferative diseases involves administration of chemotherapeutic agents, typically systemic administration. Typically chemotherapeutic agents are cytotoxic agents that act by inhibiting proliferation or other metabolic processes, so that actively proliferating and growing cells will be targeted by the agent. Such targeting, however, is not highly specific, and the side-effects are often devastating.
- Thus, a goal in pharmacology is the design of specific agents that act with high specific activity on targeted cells or tissues. This aim is of particular importance, for example, in the design of agents for treatments of diseases, such as proliferative diseases, including neoplastic disease, and diseases of viral origin, in which the ratio of toxic dose to therapeutic dose is generally close to one and the dosage must be restricted. Numerous approaches to achieving this goal have been developed. Among these are the use of conjugates that contain a targeting agent, such as an antibody and/or growth factor, and a therapeutic agent, that act on specific cells; the use of antisense technology that is targeted to specific genes and/or proteins; the use of genetic therapy to provide, for example, correct copies of defective genes or pharmaceutically active compounds, and the use of toxins that are relatively non-toxic unless delivered intracellularly. Thus far success has been limited. There are only a limited number and type of potential targeting agents, and the specificity of such agents is optimal.
- Hence there is a need to develop means for delivery of therapeutic agents to targeted cells and tissues. Therefore, it is an object herein, among others, to provide methods and compounds for targeted delivery of therapeutic agents.
- Provided herein are compounds and methods for targeted delivery of therapeutic agents. The compounds are conjugates that contain a peptidic substrate for a cell surface protease, or a soluble, shed or released form thereof, and an agent that upon cleavage by the protease is a therapeutic agent or in a form that can be activated by the targeted cell or tissue or in the localter thereof. The agents include therapeutic agents, such as a cytotoxic agents, drugs, therapeutic nucleic acid moleulces, and diagnostic agents, such as labelled moieties and imaging agents. The cell surface proteases are proteases located at a cell surface and, include, but are not limited to, membrane-bound proteases such as membrane-bound serine proteases (SPs), including, for example, proteases designated MTSPs and endotheliases. Also contemplated are proteases that are located at the cell surface by virtue of a specific binding interaction with a receptor therefor. Included among such proteases is urokinase plasminogen activator (u-PA; see, e.g., Hung (1984) Adv. Exp. Med. Biol. 172:281-293; Cheng et al. (1989)Gene 69:357-363) bound to urokinase plasminogen activator receptor (u-PAR).
- The conjugates contain one or more substrates for one or a plurality of cell surface proteases linked either directly or via a linker to a targeted agent, including a therapeutic agent, such as a cytotoxic agent. The conjugates provided herein contain the following components: (peptidic substrate)s, (linker)q, and (targeted agent)t in which: at least one peptidic substrate moiety is linked with or without a linker (L) to at least one therapeutic agent, s is 1 or more and each substrate is the same or different, and is typically is between 1 and 6, generally 1, 2 or 3; q is 0 or more as long as cell surface protease(s) cleaves the peptidic substrate(s) and releases active therapeutic agent or, releases the agent in a form that is converted by the cell, tissue or surrounding environment to an active form, q is 0 to t, generally 1 to 4; t is 1 or more, generally 1 or 2 and each targeted agent are the same or different; linker refers to any linker; and the targeted agent is any agent, typically a therapeutic agent, such as a cytotoxic agent, a nucleic acid, a diagnostic agent, such as an imaging agent or labeled moiety, or a drug, including, but not limited to, anti-tumor, anti-cancer, anti-angiogenic, pro-apoptotic and anti-mitotic agents or treatments.
- The therapeutic agents include any biologically active molecule. These agents include toxins, cytokines and lymphokines, growth factors, nucleic acid molecules, such as antisense nucleic acid, dsRNA, and DNA molecules. The therapeutic agents include those that are active intracellularly, such as cytotoxins, or extracellularly, such as modulators of the activity of extracellular receptors. When in the conjugates the therapeutic agents are substantially inactive, and when cleaved are released in active form or in a form that can be activated by the targeted cell or tissue or environment thereof.
- In an exemplary embodiment, the conjugates for use in the methods and compositions provided herein can be represented by the formula:
- (peptidel)s-(linker)q-(therapeutic agent)t
- or a derivative thereof, where peptidel is a peptidic substrate for a cell surface protease; s is greater than or equal to 1, or is 1 to 6, or is 1 or 2, or is 1; linker is any linker; q is greater than or equal to 0, or is 0 to 4, or is 0 or 1; the therapeutic agent is, for example, a cytotoxic agent, including, but not limited to, an anti-tumor, anti-angiogenic, anti-cancer, pro-apoptotic and anti-mitotic agents; and t is 1 or more, or is 1 or 2. In these conjugates, the therapeutic agent is covalently attached, optionally via a linker L, to either the C-terminus or the N-terminus of the peptidic substrate.
- In certain embodiments, peptidel is a substrate for a cell surface protease whereby, upon action of the protease, the conjugate, which is substantially inactive, is cleaved at a point on the peptidic substrate chain to release a compound of the formula:
- (peptidea)s-(linker)q-(therapeutic agent)t
- or a derivative thereof, that exhibits therapeutic activity in vitro and/or in vivo. In these conjugates, the therapeutic agent is, for example, a cytotoxic agent, and peptidea is a truncated version of peptidel resulting from cleavage at the P1-P1′ bond.
- The conjugates can be used to target and deliver the targeted agents to specific cells, and hence can be used for the treatment any diseases that are associated with cells or tissues that express a cell surface protease, including cell-associated and cell-localized proteases. The cells on which or near which such proteases are expressed are not necessarily involved in the disease or disease process, but are present and can serve to present the protease, which cleaves the targeted conjugate.
- Methods of treatment of diseases associated with cells or tissues that express a cell surface protease, including cell-associated and cell-localized proteases. The diseases include, but not limited to, proliferative diseases, autoimmune diseases, infectious diseases and inflammatory diseases. For example, diseases include e, but are not limited to, rheumatoid arthritis, lupus, multiple sclerosis, psoriasis, diabetic retinopathies, other ocular disorders, including recurrence of pterygii, scarring excimer laser surgery and glaucoma filtering surgery, various disorders of the anterior eye, cardiovascular disorders, restenosis, chronic inflammatory diseases, wounds, circulatory disorders, crest syndromes, bacterial infections, viral diseases, including AIDS, dermatological disorders, and cancer, including solid neoplasms and vascular tumors, including, but are not limited to, lung, colon, esophageal, breast, ovarian and prostate cancers.
- Also provided are methods for identifying proteases to target conjugates for treatment of diseases. The methods involve identifying cell-surface protease-associated disease by identifying a cell involved in the disease process or a cell in the vicinity of the cell involved in the disease process; and identifying a cell surface protease on the cell. Conjugates that target such proteases as provided herein can then be prepared.
- FIGS.1-5 provide in vitro CT50 (time for 50% cleavage) (min) for exemplary conjugates provided herein: A=0.1-25 min; B=25-100 min; C=100-250 min; D=>250 min.
- FIG. 1 shows exemplary doxorubicin conjugates provided herein and in vitro CT50 (min) data for cleavage of the conjugates by MTSP1.
- FIG. 2 shows exemplary doxorubicin conjugates provided herein and in vitro CT50 (min) data for cleavage of the conjugates by u-PA.
- FIG. 3 shows exemplary taxol conjugates provided herein and in vitro CT50 (min) data for cleavage of the conjugates by MTSP1.
- FIG. 4 shows exemplary taxol conjugates provided herein and in vitro CT50 (min) data for cleavage of the conjugates by u-PA.
- FIG. 5 shows exemplary doxorubicin and taxol conjugates provided herein and in vitro CT50 (min) data for cleavage of the conjugates by ET1 (endotheliase 1).
- Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of skill in the art to which the invention(s) belong. All patents, patent applications, published applications and publications, Genbank sequences, websites and other published materials referred to throughout the entire disclosure herein, unless noted otherwise, are incorporated by reference in their entirety. In the event that there are a plurality of definitions for terms herein, those in this section prevail. Where reference is made to a URL or other such indentifier or address, it understood that such identifiers can change and particular information on the internet can come and go, but equivalent information can be found by searching the internet. Reference thereto evidences the availability and public dissemination of such information.
- As used herein, a targeted agent is any agent intended for targeted delivery and includes therapeutic agents and diagnostic agents and any other agent intended for targeted delivery.
- As used herein, targeted delivery means delivery to a selected cell or tissue that expresses a protease that releases the targeted agent. Such delivery does not have to be exclusively to such selected cell or tissue, but must include it, and generally deliveries higher amounts to such selected cells or tissues. Delivery includes introduction into a cell or tissue or binding to the cell or tissue or release in the vicinity of the cell or tissue. For example, in some instances, a tumor induces production of proteases, receptors, co-factors or substrates asssociated with the stroma; delivery, thus, includes targeting such induced stromal activities, such as proteases, receptors and/or enzyme co-factors, in invading cells or cells in the tumor that is targeted.
- As used herein, therapeutic index is the ratio of LD50/ED50.
- As used herein, a therapeutic agent is any drug or other agent that is intended for delivery to a targeted cell or tissue, such as proliferating cells, including tumor cells and cells involved in a proliferative, typically an undesirable, response. Therapeutic agents, include, but are not limited to, anti-cancer agents, anti-angiogenic agents, pro-apoptotic agents, anti-mitotic growth factors, cytokines, such as tumor necrosis factors and interleukins, and cytotoxic agents and other such agents as described herein and known to those of skill in the art. Therapeutic agents include those that are active upon internalization and also those that act extracellularly, such modulators of the activities of certain cell surface receptors, such as G proteins that transduce extracellular signals.
- As used herein, an inactive therapeutic agent is a therapeutic agent that is conjugated to a peptide and thereby, either by virtue of conformational changes or size or other factors such as steric hinderance does not exhibit any or exhibits substantially reduced activity compared to the released active therapeutic agent. For example, conjugated doxorubicin is not toxic to cells until it is released from the conjugate in a form that can enter the cell. Upon cleavage of the agent from the conjugate it is in active form or in a form that is further processed by one or a plurality of steps, including enzymatically or chemically, in or on the cell, into an active form.
- As used herein, an active therapeutic agent is a therapeutic agent that has been released from the conjugate by cleavage of the peptidic substrate portion of the conjugate. The active therapeutic agent is by virtue of cleavage able to exhibit its intended activity, typically by entering the cell. When conjugated the therapeutic agents have reduced or no activity as therapeutic agents, and upon cleavage are released in the vicinity of a cell.
- As used herein, an anti-cancer agent (used interchangeably with “anti-tumor or anti-neoplasm agent”) refers to any agents used in the treatment of cancer. These include any agents, when used alone or in combination with other compounds, that can alleviate, reduce, ameliorate, prevent, or place or maintain in a state of remission of clinical symptoms or diagnostic markers associated with neoplasm, tumor or cancer, and can be used in methods, combinations and compositions provided herein. Non-limiting examples of anti-neoplasm agents include anti-angiogenic agents, alkylating agents, antimetabolite, certain natural products, platinum coordination complexes, anthracenediones, substituted ureas, methylhydrazine derivatives, adrenocortical suppressants, certain hormones, antagonists and anti-cancer polysaccharides.
- As used herein, substantially inactive with reference to the conjugated thereapeutic agent means at least 1%, generally 10, 20, 30, 50, 60, 70, 80 or 90 or 100% inactive compared to the unconjugated therapeutic agent in a standard or art-recognized assays, such as in vitro or in vivo assays, that assess the therapeutic activity of the agent.
- As used herein, a targeted cell or tissue refers to the cells or tissues that include cell surface proteases that cleave the conjugates. The cells or tissues can be involved in the disease or can be present at the disease loci or locus by virtue of participation in the disease process or merely serendipitously.
- As used herein, angiogenesis is intended to broadly encompass the totality of processes directly or indirectly involved in the establishment and maintenance of new vasculature (neovascularization), including, but not limited to, neovascularization associated with tumors.
- As used herein, anti-angiogenic treatment or agent refers to any therapeutic regimen and compound, that, when used alone or in combination with other treatment or compounds, can alleviate, reduce, ameliorate, prevent, or place or maintain in a state of remission, one or more clinical symptoms or diagnostic markers associated with undesired and/or uncontrolled angiogenesis. Thus, for purposes herein an anti-angiogenic agent refers to an agent that inhibits the establishment or maintenance of vasculature. Such agents include, but are not limited to, anti-tumor agents, and agents for treatments of other disorders associated with undesirable angiogenesis, such as diabetic retinopathies, hyperproliferative disorders and others.
- As used herein, non-anti-angiogenic anti-tumor agents refer to anti-tumor agents that do not act primarily by inhibiting angiogenesis. Whether anti-tumor agents act primarily by inhibiting angiogenesis can be determined using the assays provided herein, or using other assays well known to those of skill in the art.
- As used herein, undesired and/or uncontrolled angiogenesis refers to pathological angiogenesis wherein the influence of angiogenesis stimulators outweighs the influence of angiogenesis inhibitors. As used herein, deficient angiogenesis refers to pathological angiogenesis associated with disorders where there is a defect in normal angiogenesis resulting in aberrant angiogenesis or an absence or substantial reduction in angiogenesis.
- As used herein, a cell surface protease is any protease that is located on or at a cell surface and/or proteases that are located at the cell surface by virtue of a specific binding interaction with a receptor therefor, or that is localized at or near or associated with the cell surface. An exemplary protease located at the cell surface by virtue of a specific binding interaction with a receptor therefor is urokinase plasminogen activator (u-PA) bound to urokinase plasminogen activator receptor (u-PAR). Hence cell surface proteases contemplated herein include cell surface-associated proteases. It also includes all forms thereof that can be circulating or inside a cell. To be categorized as a cell surface protease, there must be at least one form thereof that is located (i.e. on the surfaces such as transmembrane protease or bound to receptor therefor) on the surface of a cell at some point in its cycle. Cell surface protease include serine proteases, such as, but are not limited to, the transmembrane serine protease (MTSPs) and endotheliases and urokinases.
- As used herein, a serine protease (SP) refers to a diverse family of proteases in which a serine residue is involved in the hydrolysis of proteins or peptides. The serine residue can be part of the catalytic triad mechanism, which includes a serine, a histidine and an aspartic acid in the catalysis, or be part of the hydroxyl/ε-amine or hydroxyl/α-amine catalytic dyad mechanism, which involves a serine and a lysine in the catalysis. Of particular interest are SPs of mammalian, including human, origin. Those of skill in this art recognize that, in general, single amino acid substitutions in non-essential regions of a polypeptide do not substantially alter biological activity (see, e.g., Watson et al. (1987)Molecular Biology of the Gene, 4th Edition, The Bejacmin/Cummings Pub. co., p.224).
- As used herein shed, soluble and released forms of cell surface proteases are contemplated. Such forms include, for example, forms found in serum upon proteolytic degradation or other removal of the extracellular portion of membrane bound protease, and splice variants that do not include a transmembrane domain.
- As shown herein, the protease activity of cell surface proteases and proteases associated with cells can be exploited to provide a means to concentrate therapeutic agents, such as cytotoxic agents, near such cells by providing conjugates that are activated upon cleavage by such enzymes. Such conjugates, upon the action of a cell surface protease or cell-associate protease, release the therapeutic agent, such as a cytotoxic agent, or a derivative thereof that can be converted to a therapeutic agent, locally at the site of action. As noted above, the substrates are designed to be substrates of targeted proteases that are expressed or are active on the surfaces of cells, such as tumor cells or endothelial cells, involved in or present at the site(s) or locus or loci of the disease. By virtue of specific expression, localization or activation of such proteases or the presence of receptors, substrates or enzyme co-factors therefor, administration of the conjugates provided herein permits targeting of therapeutic agents to such cells. Upon contacting with the proteases, active therapeutic agents are released in the immediate vicinity of the targeted cells. For example, specific profiles of some of the MTSPs are as follows.
- As used herein, “transmembrane serine protease (MTSP)” refers to a family of transmembrane serine proteases that share common structural features as described herein (see, also Hooper et al. (2001)J. Biol. Chem.276:857-860). Thus, reference, for example, to “MTSP” encompasses all proteins encoded by the MTSP genes, including but are not limited to: MTSP1, MTSP3, MTSP4, MTSP6, MTSP7, MTSP9, MTSP10, MTSP12, MTSP20, MTSP22 and MTSP25 or an equivalent molecule obtained from any other source or that has been prepared synthetically or that exhibits the same activity. Other MTSPs include, but are not limited to, corin, enteropeptidase, human airway trypsin-like protease (HAT), TMPRSS2 and TMPRSS4. The MTSPs described herein can be used to identify other MTSPs. Methods for isolating nucleic acid encoding other MTSPs, including nucleic acid molecules encoding full-length molecules and splice variants and MTSPs from species, such as cows, sheep, goats, pigs, horses, primates, including chimpanzees and gorillas, rodents, dogs, cats and other species of interest, such as domesticated animals, farm and zoo animals are known to those of skill in the art and are outlined herein. The nucleic acid molecules described herein including those set forth in SEQ IDs can be used to obtain nucleic acid molecules encoding full-length MTSP polypeptides from human sources or from other species, such as by screening appropriate libraries using the nucleic acid molecules or selected primers or probes based thereon.
- Sequences of encoding nucleic acid molecules and the encoded amino acid sequences of exemplary MTSPs and/or domains thereof are set forth in SEQ ID Nos. 1-45, 269-270 and 272-276. The term also encompasses MTSPs with amino acid substitutions that do not substantially alter activity of each member and also encompasses polyeptides encoded by splice variants thereof. Hence, encompassed are MTSPs with amino acid substitutions such that the resulting polypeptide retains at least 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% of the proteolytic activity of the unaltered polypeptide, and also encompasses MTSPs encoded by splice variants thereof and MTSPs encoded by allelic variants, such as single nucleotide polymorphisms (SNPs). Suitable substitutions, including, although not necessarily, conservative substitutions of amino acids, are known to those of skill in this art and can be made without eliminating the biological activity, such as the catalytic activity, of the resulting molecule. MTSPs include those of animal, such as mammalian, including human, origin.
- As used herein, a “protease domain of an MTSP” refers to an extracellular protease domain of an MTSP that exhibits proteolytic activity and shares homology and structural features with the chymotryp-sin/trypsin family protease domains. Hence it is at least the minimal portion of the domain that exhibits proteolytic activity as assessed by standard in vitro assays. Contemplated herein are such protease domains and catalytically active portions thereof.
- Exemplary MTSP polypeptides, with the protease domains indicated, are set forth in SEQ ID Nos. 1-45, 269-270 and 272-276, and including smaller portions thereof that retain or exhibit protease activity. The protease domains vary in size and constitution, including insertions and deletions in surface loops. They retain conserved structure, including at least one of the active site triad, primary specificity pocket, oxyanion hole and/or other features of serine protease domains of proteases. Thus, for purposes herein, the protease domain is a portion of a MTSP, as defined herein, and is homologous to a domain of other MTSPs. MTSPs include, MTSP1, MTSP3, MTSP4, MTSP6, MTSP7, MTSP9, MTSP10, MTSP12, MTSP20, MTSP22 and MTSP25 (see SEQ ID Nos. 1-19, 42-45, 269-270 and 272-276; see, also International PCT application No. WO 02/00860 (see SEQ ID Nos. 38 and 97 therein, which provide an MTSP12 variant); corin (SEQ ID Nos. 28 and 29), enteropeptidase (SEQ ID Nos. 30 and 31) human airway trypsin-like protease (HAT) (SEQ ID Nos. 32 and 33), hepsin (SEQ ID Nos. 34 and 35), TMPRSS2 (SEQ ID Nos. 36 and 37) and TMPRSS4 (SEQ ID Nos. 38 and 39). As with the larger class of enzymes of the chymotrypsin (S1) fold (see, e.g., Internet accessible MEROPS data base), the MTSPs protease domains share a high degree of amino acid sequence identity. The His, Asp and Ser residues necessary for activity are present in conserved motifs. In those that are activated by cleavage, the activation site, which results in the N-terminus of second chain in the two chain forms has a conserved motif and readily can be identified (see, e.g., amino acids 801-806, SEQ ID No. 29, amino acids 406-410, SEQ ID No. 31; amino acids 186-190, SEQ ID No. 33; amino acids 161-166, SEQ ID No. 35; amino acids 255-259, SEQ ID No. 37; amino acids 190-194, SEQ ID No. 39 and other as known to those of skill and the art and/or as described herein).
- For example, with reference to MTSP10 (see SEQ ID Nos. 44 and 45), there disulfide bonds as follows: C488-C504, C587-C653; C619-C632; C643-C673 (see SEQ ID Nos. 44 and 45) (chymotrypsin numbering 42 to 58; 136-201; 168-182 and 191-220). Disulfide bonds form between the Cys residues C573-C296 to link the protease domain to another domain so that upon activation cleavage (between residues R462 and I463 of SEQ ID No. 45) the resulting polypeptide is a two chain molecule. The C573 (SEQ ID NO. 45 is a free Cys in a single chain form of the protease domain. As noted the protease also can be provided as a two chain molecule. Single chain and two chain forms are proteolytically active. A two chain form is produced by bonding, typically between the C573 and a Cys outside the protease domain, such as Cys296. Upon activation cleavage the disulfide bond remains resulting in a two chain polypeptide. The size of chain “A” is a function the starting length of the polypeptide prior to activation cleavage between the R462 and I463. Any length polypeptide that includes the protease domain (residues 463-692 of SEQ ID No. 45) or catalytically active fragments thereof, is contemplated herein. Two chain forms include at least the protease domain a polypeptide from C296 up to and including C573.
- As used herein, a two-chain form of the protease domain refers to a two-chain form that is formed from a single chain form of the protease in which the Cys pairing between, e.g., a Cys outside the protease domain such as, for example Cys573 (SEQ ID No. 45 for MTSP), which links the protease domain to the remainder of the polypeptide, the “A” chain. A two chain protease domain form refers to any form in which the “remainder of the polypeptide”, i.e., “A” chain, is shortened and includes a Cys from outside the protease domain.
- As used herein, the catalytically active domain of an MTSP refers to the protease domain. Reference to the protease domain of an MTSP generally refers to a single chain form of the protein. If the two-chain form or both forms is intended, it is so-specified. The zymogen form of each protein is a single chain, which is converted to the active two or multi chain form by activation cleavage. By active form is meant a form active in vivo or in vitro.
- As used herein, activation cleavage refers to the cleavage of the protease at the N-terminus of the protease domain (generally between an R and I or V in the full-length protein. By virtue of the Cys-Cys pairing between a Cys outside the protease domain and a Cys in the protease domain (see, e.g., Cys573 SEQ ID No. 45, upon cleavage the resulting polypeptide has two chains (“A” chain and the “B” chain, which is the protease domain of an MTSP). Cleavage can be effected by another protease or autocatalytically. The conjugates provided herein advantageously contain sites that are recognized by the active cell surface protease (or cell-associated protease) and are cleaved thereby to release active or an inactive prodrug form of a therapeutic agent.
- As used herein an MTSP1, whenever referenced herein, includes at least one or all of or any combination of:
- a polypeptide encoded by the sequence of nucleotides set forth in SEQ ID No. 1 or 40;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in SEQ ID No. 1 or 40;
- a polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 2 or 41;
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 2 or 41; and/or
- a polypeptide encoded by a splice variant of the MTSP1 set forth in SEQ ID No. 1 or 40.
- The MTSP1 can be from any animal, particularly a mammal, and includes but is not limited to, humans, rodents, fowl, ruminants and other animals. The full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form. MTSP1 also is referred to TADG-15 and matriptase. As described below, the protein originally designated matriptase appears to be an MTSP1 splice variant or processed product.
- As used herein an MTSP3, whenever referenced herein, includes at least one or all of or any combination of:
- a polypeptide encoded by the sequence of nucleotides set forth in SEQ ID No. 3;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in SEQ ID No. 3;
- a polypeptide that comprises the sequence of amino acids set forth as amino acids 205-437 of SEQ ID No. 4;
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 4; and/or
- a polypeptide encoded by a splice variant of the MTSP3 set forth in SEQ ID Nos. 3 and 4.
- The MTSP3 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals. The full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- As used herein an MTSP4, whenever referenced herein, includes at least one or all of or any combination of:
- a polypeptide encoded by the sequence of nucleotides set forth in any of SEQ ID No. 5, 7 or 9;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in any of SEQ ID Nos. 5, 7 or 9;
- a polypeptide that comprises the sequence of amino acids set forth in any of SEQ ID Nos. 6, 8 or 10;
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 6, 8 or 10; and/or
- a polypeptide encoded by a splice variant of the MTSP4s set forth in SEQ ID Nos. 7-10.
- The MTSP4 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals. The full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- As used herein an MTSP6, whenever referenced herein, includes at least one or all of or any combination of:
- a polypeptide encoded by the sequence of nucleotides set forth in any of SEQ ID No. 11;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in any of SEQ ID Nos. 11;
- a polypeptide that comprises the sequence of amino acids set forth in any of SEQ ID No. 12;
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 12; and/or
- a polypeptide encoded by a splice variant of the MTSP6 set forth in SEQ ID No. 12.
- The MTSP6 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals. The full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form. Of particular interest herein is the MTSP6 of SEQ ID No. 12.
- As used herein an MTSP7, whenever referenced herein, includes at least one or all of or any combination of:
- a polypeptide encoded by the sequence of nucleotides set forth in SEQ ID No. 13;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in SEQ ID No. 13;
- a polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 13;
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 14; and/or
- a polypeptide encoded by a splice variant of the MTSP7 set forth in SEQ ID No. 13.
- The MTSP7 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals. The full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- As used herein an MTSP9, whenever referenced herein, includes at least one or all of or any combination of:
- a polypeptide encoded by the sequence of nucleotides set forth in SEQ ID No. 17 or SEQ ID No. 42;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in SEQ ID No. 17 or 42;
- a polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 18 or 43;
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 18 or 270; and/or
- a polypeptide encoded by a splice variant of the MTSP9 set forth in SEQ ID No. 17.
- The MTSP9 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals. The full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- As used herein an MTSP10, whenever referenced herein, includes at least one or all of or any combination of:
- a polypeptide encoded by the sequence of nucleotides set forth in SEQ ID No. 44;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in SEQ ID No. 44;
- a polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 45;
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 45; and/or
- a polypeptide encoded by a splice variant of the MTSP10 set forth in SEQ ID No. 44.
- The MTSP10 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals. The full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form.
- MTSP10 polypeptides, including, but not limited to splice variants thereof, and nucleic acids encoding MTSPs, and domains, derivatives and analogs thereof are provided herein. Single chain protease domains that have an N-terminus functionally equivalent to that generated by activation of the zymogen form of MTSP10 are also provided. The cleavage site for the protease domain of MTSP10 is between amino acid R and amino acids I (R↓IIGGT) (residues 462-467 SEQ ID No. 45).
- As used herein an MTSP12, whenever referenced herein, includes at least one or all of or any combination of: SEQ ID No. 19 and 20
- a polypeptide encoded by the sequence of nucleotides set forth in SEQ ID No. 19 or by a sequence of nucleotides that includes nucleotides that encode the sequence of amino acids set forth in SEQ ID No. 20;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in is set forth as SEQ ID No. 19;
- a polypeptide that includes the sequence of amino acids set forth in SEQ ID No. 20 or a catalytically active portion thereof;
- a polypeptide that includes a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 20; and/or
- a polypeptide encoded by a splice variant of the MTSP12 that includes the sequence of amino acids set forth in SEQ ID No. 20.
- In particular, the MTSP12 polypeptide, with the protease domains as indicated in SEQ ID Nos. 19 and 20, is provided. The polypeptide is a single or multi-chain polypeptide. A protease domain of an MTSP12, whenever referenced herein, includes at least one or all of or any combination of or a catalytically active portion of:
- a polypeptide that includes the sequence of amino acids set forth in SEQ ID No. 20 or a catalytically active portion thereof but that does not include the sequence of amino acids set forth in SEQ ID No. 271;
- a polypeptide that includes the sequence of amino acids set forth in SEQ ID No. 272 or a catalytically active fragment thereof;
- a polyeptide containing amino acids 237 to 456 of SEQ ID No. 6, a polypeptide containing
amino aicds 538 to 765 of SEQ ID No. 20, and a polypeptide containing amino acids 861 to 1087 of SEQ ID No. 20, but that does not include the sequence of amino acids set forth in SEQ ID No. 271; - a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to a sequence of nucleotides that encodes any of the polypeptides of a)-c);
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in SEQ ID No. 20 but that does not encode the sequence of amino acids set forth in SEQ ID No. 271;
- a polypeptide that includes a sequence of amino acids having at least about 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 20;
- a polypeptide that includes a sequence of amino acids having at least about 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids of the polypeptides of a)-e);
- a polypeptide encoded by a splice variant of a sequence of nucleotides that encodes an MTSP12 of any of the above.
- Smaller portions thereof that retain protease activity are also provided. The MTSP12 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals. The full-length zymogen or two-chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two-chain activated form, or a single chain form. MTSP12 also includes the variant described International PCT application No. WO 02/00860 (see SEQ ID Nos. 38 and 97 therein).
- As used herein an MTSP20, whenever referenced herein, includes at least one or all of or any combination of:
- a polypeptide encoded by the sequence of nucleotides set forth in SEQ ID No. 273;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in SEQ ID No. 273;
- a polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 273;
- a polypetide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 274; and/or a polypeptide encoded by a splice variant of the MTSP20 encoded by the sequence of nucleotides that includes the sequence set forth in SEQ ID No. 273.
- The MTSP20 may be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals. The full length zymogen or two-chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two-chain activated form, or a single chain form.
- As used herein an MTSP22, whenever referenced herein, includes at least one or all of or any combination of:
- a polypeptide encoded by the sequence of nucleotides set forth in SEQ ID No. 275;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in SEQ ID No. 275;
- a polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 276;
- a polypetide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 276; and/or
- a polypeptide encoded by a splice variant of the MTSP22 set forth in SEQ ID No. 275.
- The MTSP22 may be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals. The full length zymogen or two-chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two-chain activated form, or a single chain form.
- As used herein an MTSP25, whenever referenced herein, includes at least one or all of or any combination of:
- a polypeptide encoded by the sequence of nucleotides set forth in SEQ ID No. 269;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in SEQ ID No. 269;
- a polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 270;
- a polypetide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 270; and/or
- a polypeptide encoded by a splice variant of the MTSP25 set forth in SEQ ID No. 269.
- The MTSP25 may be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals. The full length zymogen or two-chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two-chain activated form, or a single chain form.
- As used herein, a human protein is one encoded by nucleic acid present in the genome of a human, including all allelic variants and conservative variations as long as they are not variants found in other mammals.
- As used herein, not substantially cleaved by plasmin or prostate specific antigen (PSA) (or other non-cell surface-associated protease), means in comparable in vitro assays (under optimal conditions for each enzyme) in which the activity of a targeted cell surface membrane protease or catalytically active portion of the activity of the protease domain (or a catalytically active form thereof) of prostate specific antigen (PSA) or plasmin for cleavage of the conjugate is compared, the relative activity is greater than at least 2:1, 3:1, 4:1, 5:1, 10:1, 50:1 or 100:1.
- As used herein, activity refers to the ratio kcat/Km, where kcat is the rate of catalytic turnover for a particular enzyme, and Km is the Michaelis constant for the binding of the substrate.
- As used herein, a “nucleic acid encoding a protease domain or catalytically active portion of a MTSP” shall be construed as referring to a nucleic acid encoding only the recited single chain protease domain or active portion thereof, and not the other contiguous portions of the MTSP as a continuous sequence.
- As used herein, a CUB domain is a motif that mediates protein-protein interactions in complement components C1r/C1s and has also been identified in various proteins involved in developmental processes.
- As used herein, a zymogen is an enzymatically inactive protein (i.e, typically, but not necessarily, less than 1% of active form) that is converted to a proteolytic enzyme by the action of an activator, including by autoactivation. Inactive means less active than the form those of skill in the art consider to be the active form of the enzyme. The ratio of activity of a zymogen to the activated form varies from enzyme-to-enzyme.
- As used herein, “disease or disorder” refers to a pathological condition in an organism resulting from, e.g., infection or genetic defect, and characterized by identifiable symptoms. The diseases contemplated for treatment herein are any for which a cell surface protease, including a cell-localized or cell-associated protease is asssociated with a targeted cell or tissue involved in the disease or disease process. Such association can be because the protease is involved in the disease or is serendipitously associated with cells involved with the disease. These diseases herein are called cell surface protease-associated diseases. Hence, to treat th disease a cellsurface protease is identified that is expressed on cells associated with the disorder, such as, for example, immune cells for treating inflammatory diseases, and virally infected cells for treating viral diseases. The conjugate is designed as described herein for cleavage by the selected protease.
- As used herein, neoplasm (neoplasia) refers to abnormal new growth, and thus means the same as tumor, which can be benign or malignant. Unlike hyperplasia, neoplastic proliferation persists even in the absence of the original stimulus.
- As used herein, neoplastic disease refers to any disorder involving cancer, including tumor development, growth, metastasis and progression.
- As used herein, cancer refers to a general term for diseases caused by any type of malignant tumor.
- As used herein, malignant, as applied to tumors, refers to primary tumors that have the capacity of metastasis with loss of growth control and positional control.
- As used herein, endotheliase refers to a mammalian protein, including human protein, that has a transmembrane domain and is expressed or active on the surface of endothelial cells and includes a protease domain, particularly an extracellular protease domain, and is generally a serine protease (see, also U.S. application Ser. No. 09/717,473 and International PCT application No. WO 01/36604). Thus, reference, for example, to endotheliase encompasses all proteins encoded by the endotheliase gene family, or an equivalent molecule obtained from any other source or that has been prepared synthetically or that exhibits the same activity. The endotheliase gene family are transmembrane proteases expressed or active in endothelial cells. These proteases include serine proteases. These include proteins that have these features and also include a protease domain that exhibits sequence homology to the
endotheliases - As used herein an
endotheliase 1, whenever referenced herein, includes at least one or all of or any combination of: - a polypeptide encoded by the sequence of nucleotides set forth in SEQ ID No. 21;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in SEQ ID No. 21;
- a polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 22;
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 22; and/or
- a polypeptide encoded by a splice variant of a nucleic acid molecule that encodes a protein containing the polypeptide set forth in SEQ ID No. 22.
- The
endotheliase 1 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals. The full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form. - As used herein an
endotheliase 2, whenever referenced herein, includes at least one or all of or any combination of: - a polypeptide encoded by the sequence of nucleotides set forth in SEQ ID No. 23 or 25;
- a polypeptide encoded by a sequence of nucleotides that hybridizes under conditions of low, moderate or high stringency to the sequence of nucleotides set forth in SEQ ID No. 23 or 25;
- a polypeptide that comprises the sequence of amino acids set forth in SEQ ID No. 24 or 26;
- a polypeptide that comprises a sequence of amino acids having at least about 40%, 60%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with the sequence of amino acids set forth in SEQ ID No. 24 or 26; and/or
- a polypeptide encoded by a splice variant of a nucleic acid set forth in SEQ ID No. 23 or 25.
- The
endotheliase 2 can be from any animal, particularly a mammal, and includes but are not limited to, humans, rodents, fowl, ruminants and other animals. The full length zymogen or two chain activated form is contemplated or any domain thereof, including the protease domain, which can be a two chain activated form, or a single chain form. - As used herein, the protease domain of an endotheliase refers to the polypeptide portion of the endotheliase that is the extracellular portion that exhibits protease activity. The protease domain is a polypeptide that includes at least the minimum number of amino acids, generally more than 50 or 100, required for protease activity. Protease activity can be assessed empirically, such as by testing the polypeptide for its ability to act as a protease. Assays, such as those described in the EXAMPLES, with the exception that a known endotheliase substrate is employed in place of the test compounds, can be used to assess protease activity. Furthermore, since proteases, particularly serine proteases, have characteristic structures and sequences or motifs, the protease domain can be readily identified by such structure and sequence or motif.
- As used herein, a portion of protease domain of endotheliase refers to a portion of protease domain of endotheliase that is located within or is the extracellular domain of an endotheliase and exhibits serine proteolytic activity. Hence, it is at least the minimal portion of the extracellular domain that exhibits proteolytic activity as assessed by standard assays. An exemplary protease domain of an endotheliase is set forth in SEQ ID No. 22 and as amino acids 321-688 and 321-562 of SEQ ID Nos. 24 and 26, respectively. Smaller portions thereof that retain protease activity are contemplated. The protease domains vary in size and constitution, including insertions and deletions in surface loops. Such domains exhibit conserved structure, including at least one structural feature, such as the active site triad, primary specificity pocket, oxyanion hole and/or other features of serine protease domains of proteases. Thus, for purposes herein, the protease domain is a portion of an endotheliase, as defined herein, but is homologous in terms of structural features and retention of sequence of similarity or homology the protease domain of chymotrypsin or trypsin.
- As used herein, homologous means about greater than about 25%, 40%, 60%, 80%, 90%, 95%, 98% or greater sequence identity. By sequence identity, the number of conserved amino acids as determined by standard alignment algorithms programs, and used with default gap penalties established by each supplier. Also homology can be assessed by conserved nucleic acid sequence, which includes anything that hybridizes under at least low stringency conditions and encodes the domain. Similarly, nucleic acid sequence alignment programs are commercially available (DNAStar “MegAlign” program (Madison, Wis.) and the University of Wisconsin Genetics Computer Group (UWG) “Gap” program (Madison, Wis.)). Substantially homologous nucleic acid molecules would hybridize typically at moderate stringency or at high stringency all along the length of the nucleic acid of interest. Also contemplated are nucleic acid molecules that contain degenerate codons in place of codons in the hybridizing nucleic acid molecule.
- As used herein, recitation that a polypeptide consists essentially of the protease domain means that the only endotheliase portion of the polypeptide is a protease domain or a catalytically active portion thereof. The polypeptide can optionally include additional non-endotheliase-derived sequences of amino acids.
- As used herein, domain refers to a portion of a molecule, e.g., proteins or nucleic acids, that is structurally and/or functionally distinct from other portions of the molecule.
- As used herein, an active form of a protease refers to an enzyme that catalyzes hydrolysis of proteins or peptides. Reference to a protease includes the active and zymogen or other less active form.
- As used herein, nucleic acids include DNA, RNA and analogs thereof, including peptide nucleic acids (PNA) and mixtures thereof. Nucleic acids can be single or two stranded. When referring to probes or primers, optionally labeled, with a detectable label, such as a fluorescent or radiolabel, single-stranded molecules are contemplated. Such molecules are typically of a length such that their targets are statistically unique or of low copy number (typically less than 5, generally less than 3) for probing or priming a library. Generally a probe or primer contains at least 14, 16 or 30 contiguous of sequence complementary to or identical to a gene of interest. Probes and primers can be 10, 20, 30, 50, 100 or more nucleic acids long.
- As used herein, nucleic acid encoding a fragment or portion of an endotheliase refers to a nucleic acid encoding only the recited fragment or portion of endotheliase protein, and not the other contiguous portions of the endotheliase as a continuous sequence.
- As used herein, heterologous nucleic acid is nucleic acid that, if it is DNA encodes RNA, or, if RNA, encodes proteins that generally are not normally produced in vivo by the cell in which it is expressed or that mediates or encodes mediators that alter expression of endogenous nucleic acid, such as DNA, by affecting transcription, translation, or other regulatable biochemical processes or that is located in a different locus from its normal locus. Heterologous nucleic acid is generally not endogenous to the cell into which it is introduced, but has been obtained from another cell or prepared synthetically. Generally, although not necessarily, such nucleic acid encodes RNA and proteins that are not normally produced by the cell in which it is now expressed.
- Heterologous nucleic acid, such as DNA, also be referred to as foreign nucleic acid, such as DNA. Any nucleic acid, such as DNA, that one of skill in the art would recognize or consider as heterologous or foreign to the cell in which is expressed is herein encompassed by heterologous nucleic acid; heterologous nucleic acid includes exogenously added nucleic acid that is also expressed endogenously. Examples of heterologous nucleic acid include, but are not limited to, nucleic acid that encodes traceable marker proteins, such as a protein that confers drug resistance, nucleic acid that encodes therapeutically effective substances, such as anti-cancer agents, enzymes and hormones, and nucleic acid, such as DNA, that encodes other types of proteins, such as antibodies, and RNA, such as RNA interference (RNAi) or other double-stranded RNA, and antisense RNA. Antibodies that are encoded by heterologous nucleic acid can be secreted or expressed on the surface of the cell in which the heterologous nucleic acid has been introduced.
- For example, nucleic acid can be the the targeted agent, such as the therapeutic or diagnostic agent, in the conjugate. Nucleic acids, include ds RNA use for RNA interference (RNAi) (see, e.g. Chuang et al. (2000)Proc. Natl. Acad. Sci. U.S.A. 97:4985) which is employed to inhibit the expression of a targeted gene by generating loss-of-function. Methods relating to the use of RNAi to silence genes in organisms including, mammals, C. elegans, Drosophila and plants, and humans are known (see, e.g., Fire et al. (1998) Nature 391:806-811 Fire (1999) Trends Genet. 15:358-363; Sharp (2001) Genes Dev. 15:485-490; Hammond, et al. (2001) Nature Rev. Genet.2:110-1119; Tuschl (2001) Chem. Biochem. 2:239-245; Hamilton et al. (1999) Science 286:950-952; Hammond et al. (2000) Nature 404:293-296; Zamore et al. (2000) Cell 101:25-33; Bernstein et al. (2001) Nature 409: 363-366; Elbashir et al. (2001) Genes Dev. 15:188-200; Elbashir et al. (2001) Nature 411:494-498; International PCT application No. WO 01/29058; International PCT application No. WO 99/32619). By selecting appropriate sequences, expression of dsRNA can interfere with accumulation of endogenous mRNA encoding a targeted gene product. Regions that include at least about 21 nucleotides and that are selective (i.e. whose target is unique) for the nucleic acid encoding a targeted gene product are used to prepare the RNAi.
- As used herein, genetic therapy involves the transfer of heterologous nucleic acid, such as DNA, into certain cells, target cells, of a mammal, particularly a human, with a disorder or conditions for which such therapy is sought. The nucleic acid molecules are included in a conjugate linked via a cell surface protein cleavage site. The nucleic acid, such as DNA, is introduced into the selected target cells in a manner such that the heterologous nucleic acid, such as DNA, is expressed and a therapeutic product encoded thereby is produced. Alternatively the heterologous nucleic acid, such as DNA, can in some manner mediate expression of DNA that encodes the therapeutic product, or it can encode a product, such as a peptide or RNA that in some manner mediates, directly or indirectly, expression of a therapeutic product. Genetic therapy can also be used to deliver nucleic acid encoding a gene product that replaces a defective gene or supplements a gene product produced by the mammal or the cell in which it is introduced. The introduced nucleic acid can encode a therapeutic compound, such as a growth factor inhibitor thereof, or a tumor necrosis factor or inhibitor thereof, such as a receptor therefor, that is not normally produced in the mammalian host or that is not produced in therapeutically effective amounts or at a therapeutically useful time. The heterologous nucleic acid, such as DNA, encoding the therapeutic product can be modified prior to introduction into the cells of the afflicted host in order to enhance or otherwise alter the product or expression thereof. Genetic therapy can also involve delivery of an inhibitor or repressor or other modulator of gene expression, such dsRNA or antisense or other nucleic acid molecule. The conjugates herein can be used to deliver a product, such as a nucleic acid for gene therapy.
- As used herein, a therapeutically effective product for gene therapy is a product that is encoded by heterologous nucleic acid, typically DNA, that, upon introduction of the nucleic acid into a host, a product is expressed that ameliorates or eliminates the symptoms, manifestations of an inherited or acquired disease or that cures the disease. Also included are biologically active nucleic acid molecules, such as RNAi and antisense.
- As used herein, a sequence complementary to at least a portion of an RNA, with reference to antisense oligonucleotides, means a sequence having sufficient complementarily to be able to hybridize with the RNA, generally under moderate or high stringency conditions, forming a stable duplex; in the case of double-stranded SP antisense nucleic acids, a single strand of the duplex DNA (or dsRNA) can thus be tested, or triplex formation can be assayed. The ability to hybridize depends on the degree of complementarily and the length of the antisense nucleic acid. Generally, the longer the hybridizing nucleic acid, the more base mismatches with a SP encoding RNA it can contain and still form a stable duplex (or triplex, as the case can be). One skilled in the art can ascertain a tolerable degree of mismatch by use of standard procedures to determine the melting point of the hybridized complex.
- Amino acid substitutions can be made or occur in any SPs and protease domains thereof. Amino acid substitutions include conservative substitutions, such as those set forth in Table 1, which do not eliminate proteolytic activity. As described herein, substitutions that alter properties of the proteins, such as removal of cleavage sites and other such sites are also contemplated; such substitutions are generally non-conservative, but can be readily effected by those of skill in the art.
- Suitable conservative substitutions of amino acids are known to those of skill in this art and can be made generally without altering the biological activity, for example enzymatic activity, of the resulting molecule. Also included within the definition, is the catalytically active fragment of an SP, particularly a single chain protease portion. Conservative amino acid substitutions are made, for example, in accordance with those set forth in TABLE 1 as follows:
TABLE 1 Ala (A) Gly; Ser Arg (R) Lys, Orn Asn (N) Gln; His Asp (D) Glu Cys (C) Ser Gln (Q) Asn Glu (E) Asp Gly (G) Ala; Pro His (H) Asn; Gln Ile (I) Leu; Val; Nle; Met Leu (L) Ile; Val; Nle; Met Lys (K) Arg; Gln; Glu Met (M) Leu; Tyr; Ile; Nle Phe (F) Met; Leu; Tyr, Trp Ser (S) Thr Thr (T) Ser Trp (W) Tyr; Phe Tyr (Y) Trp; Phe Val (V) Ile; Leu; Nle; Met - Other substitutions are also permissible and can be determined empirically or in accord with known conservative substitutions. For example, one or more amino acid residues within the sequence can be substituted by another amino acid of a similar polarity which acts as a functional equivalent, resulting in a silent alteration. Substitutes for an amino acid within the sequence can be selected from other members of the class to which the amino acid belongs. For example, the nonpolar (hydrophobic) amino acids include alanine, leucine, isoleucine, valine, proline, phenylalanine, tryptophan and methionine. The polar neutral amino acids include glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine. The positively charged (basic) amino acids include arginine, lysine and histidine. The negatively charged (acidic) amino acids include aspartic acid and glutamic acid.
- As used herein, the amino acids, which occur in the various amino acid sequences appearing herein, are identified according to their well-known, three-letter or one-letter abbreviations. The nucleotides, which occur in the various DNA fragments, are designated with the standard single-letter designations used routinely in the art. Other abbreviations, include: hR or hArg for homoarginine; hY or hTyr for homotyrosine; Cha for cyclohexylalanine; Amf for 4-aminomethylphenylalanine; DPL for 2-(4,6-dimethylpyrimidinyl)lysine; (imidazolyl)K for N′-(2-imidazolyl)lysine; Me2PO3-Y for 0-dimethylphosphotyrosine; O—Me—Y for O-methyltyrosine; TIC for tetrahydro-3-isoquinoline carboxylic acid; MeL for 2-keto-3-amino-5-methylhexane; DAP for 1,3-diaminopropane; TFA for trifluoroacetic acid; AA for acetic acid.
- As used herein, a splice variant refers to a variant produced by differential processing of a primary transcript of genomic DNA that results in more than one type of mRNA.
- As used herein, a probe or primer based on a nucleotide sequence disclosed herein, includes at least 10, 14, generally at least 16 or 30 or 100 contiguous sequence of nucleotides.
- As used herein, antisense polynucleotides refer to synthetic sequences of nucleotide bases complementary to mRNA or the sense strand of double-stranded DNA. Admixture of sense and antisense polynucleotides under appropriate conditions leads to the binding of the two molecules, or hybridization. When these polynucleotides bind to (hybridize with) mRNA, inhibition of protein synthesis (translation) occurs. When these polynucleotides bind to double-stranded DNA, inhibition of RNA synthesis (transcription) occurs. The resulting inhibition of translation and/or transcription leads to an inhibition of the synthesis of the protein encoded by the sense strand. Antisense nucleic acid molecules typically contain a sufficient number of nucleotides to specifically bind to a target nucleic acid, generally at least 5 contiguous nucleotides, often at least 14 or 16 or 30 contiguous nucleotides or modified nucleotides complementary to the coding portion of a nucleic acid molecule that encodes a gene of interest, for example, nucleic acid encoding a single chain protease domain of an SP.
- As used herein, an array refers to a collection of elements, such as antibodies, containing three or more members. An addressable array is one in which the members of the array are identifiable, typically by position on a solid phase support. Hence, in general the members of the array are immobilized on discrete identifiable loci on the surface of a solid phase.
- As used herein, antibody refers to an immunoglobulin, whether natural or partially or wholly synthetically produced, including any derivative thereof that retains the specific binding ability of the antibody. Hence antibody includes any protein having a binding domain that is homologous or substantially homologous to an immunoglobulin binding domain. Antibodies include members of any immunoglobulin claims, including IgG, IgM, IgA, IgD and IgE.
- As used herein, antibody fragment refers to any derivative of an antibody that is less than full-length, retaining at least a portion of the full-length antibody's specific binding ability. Examples of antibody fragments include,but are not limited to, Fab, Fab′, F(ab)2, single-chain Fvs (scFV), FV, dsFV diabody and Fd fragments. The fragment can include multiple chains linked together, such as by disulfide bridges. An antibody fragment generally contains at least about 50 amino acids and typically at least 200 amino acids.
- As used herein, an Fv antibody fragment is composed of one variable heavy domain (VH) and one variable light domain linked by noncovalent interactions.
- As used herein, a dsFV refers to an Fv with an engineered intermolecular disulfide bond, which stabilizes the VH-VL pair.
- As used herein, an F(ab)2 fragment is an antibody fragment that results from digestion of an immunoglobulin with pepsin at pH 4.0-4.5; it can be recombinantly expressed to produce the equivalent fragment.
- As used herein, Fab fragments are antibody fragments that result from digestion of an immunoglobulin with papain; they can be recombinantly expressed to produce the equivalent fragment.
- As used herein, scFVs refer to antibody fragments that contain a variable light chain (VL) and variable heavy chain (VH) covalently connected by a polypeptide linker in any order. The linker is of a length such that the two variable domains are bridged without substantial interference. Exemplarly linkers include, but are not limited to, (Gly-Ser)n residues, which can include ome Glu or Lys residues dispersed throughout, for example, to increase solubility.
- As used herein, humanized antibodies refer to antibodies that are modified to include human sequences of amino acids so that administration to a human does not provoke an immune response. Methods for preparation of such antibodies are known. For example, to produce such antibodies, the encoding nucleic acid in the hybridoma or other prokaryotic or eukaryotic cell, such as anE. coli or a CHO cell, that expresses the monoclonal antibody is altered by recombinant nucleic acid techniques to express an antibody in which the amino acid composition of the non-variable region is based on human antibodies. Computer programs have been designed to identify such non-variable regions.
- As used herein, diabodies are dimeric scFV; diabodies typically have shorter peptide linkers than scFvs, and they generally dimerize.
- As used herein, production by recombinant means by using recombinant DNA methods means the use of the well known methods of molecular biology for expressing proteins encoded by cloned DNA.
- As used herein, the term assessing is intended to include quantitative and qualitative determination in the sense of obtaining an absolute value for the activity of an SP, or a domain thereof, present in the sample, and also of obtaining an index, ratio, percentage, visual or other value indicative of the level of the activity. Assessment can be direct or indirect and the chemical species actually detected need not of course be the proteolysis product itself but can for example be a derivative thereof or some further substance.
- As used herein, biological activity refers to the in vivo activities of a compound or physiological responses that result upon in vivo administration of a compound, composition or other mixture. Biological activity, thus, encompasses therapeutic effects and pharmaceutical activity of such compounds, compositions and mixtures. Biological activities can be observed in in vitro systems designed to test or use such activities.
- As used herein, a combination refers to any association between two or among more items.
- As used herein, fluid refers to any composition that can flow. Fluids thus encompass compositions that are in the form of semi-solids, pastes, solutions, aqueous mixtures, gels, lotions, creams and other such compositions.
- As used herein, an effective amount of a compound for treating a particular disease is an amount that is sufficient to ameliorate, or in some manner reduce the symptoms associated with the disease. Such amount can be administered as a single dosage or can be administered according to a regimen, whereby it is effective. The amount can cure the disease but, typically, is administered in order to ameliorate the symptoms of the disease. Repeated administration can be required to achieve the desired amelioration of symptoms.
- As used herein, equivalent, when referring to two sequences of nucleic acids, means that the two sequences in question encode the same sequence of amino acids or equivalent proteins. When equivalent is used in referring to two proteins or peptides, it means that the two proteins or peptides have substantially the same amino acid sequence with amino acid substitutions (see, e.g., Table 1, above) that do not substantially alter the activity or function of the protein or peptide (i.e, retain at least about 1% of the activity). When equivalent refers to a property, the property does not need to be present to the same extent (e.g., two peptides can exhibit different rates of the same type of enzymatic activity), but the activities are generally substantially the same. Complementary, when referring to two nucleotide sequences, means that the two sequences of nucleotides are capable of hybridizing, typically with less than 25%, often with less than 15%, or even less than 5% or with no mismatches between opposed nucleotides. Generally the two molecules hybridize under conditions of high stringency.
- As used herein, a method for treating or preventing disease or disorder associated with undesired and/or uncontrolled angiogenesis means that the diseases or the symptoms associated with the undesired and/or uncontrolled angiogenesis are alleviated, reduced, ameliorated, prevented, placed in a state of remission, or maintained in a state of remission. It also means that the hallmarks of pathological angiogenesis are eliminated, reduced or prevented by the treatment. Non-limiting examples of the hallmarks of the pathological angiogenesis include uncontrolled degradation of the basement membrane and proximal extracellular matrix of the endothelial cells, migration, division, and organization of the endothelial cells into new functioning capillaries, and the persistence of such functioning capillaries.
- As used herein, operatively linked or operationally associated refers to the functional relationship of DNA with regulatory and effector sequences of nucleotides, such as promoters, enhancers, transcriptional and translational stop sites, and other signal sequences. For example, operative linkage of DNA to a promoter refers to the physical and functional relationship between the DNA and the promoter such that the transcription of such DNA is initiated from the promoter by an RNA polymerase that specifically recognizes, binds to and transcribes the DNA. In order to optimize expression and/or in vitro transcription, it can be necessary to remove, add or alter 5′ untranslated portions of the clones to eliminate extra, potential inappropriate alternative translation initiation (i.e., start) codons or other sequences that can interfere with or reduce expression, either at the level of transcription or translation. Alternatively, consensus ribosome binding sites (see, e.g., Kozak (1991)J. Biol. Chem. 266:19867-19870) can be inserted immediately 5′ of the start codon and can enhance expression. The desirability of (or need for) such modification can be empirically determined.
- As used herein, a promoter region or promoter element refers to a segment of DNA or RNA that controls transcription of the DNA or RNA to which it is operatively linked. The promoter region includes specific sequences that are sufficient for RNA polymerase recognition, binding and transcription initiation. This portion of the promoter region is referred to as the promoter. In addition, the promoter region includes sequences that modulate this recognition, binding and transcription initiation activity of RNA polymerase. These sequences can be cis acting or can be responsive to trans acting factors. Promoters, depending upon the nature of the regulation, can be constitutive or regulated. Exemplary promoters contemplated for use in prokaryotes include the bacteriophage T7 and T3 promoters.
- As used herein, sample refers to anything which can contain an analyte for which an analyte assay is desired. The sample can be a biological sample, such as a biological fluid or a biological tissue. Examples of biological fluids include urine, blood, plasma, serum, saliva, semen, stool, sputum, cerebral spinal fluid, tears, mucus, amniotic fluid or the like. Biological tissues are aggregates of cells, usually of a particular kind together with their intercellular substance that form one of the structural materials of a human, animal, plant, bacterial, fungal or viral structure, including connective, epithelium, muscle and nerve tissues. Examples of biological tissues also include organs, tumors, lymph nodes, arteries and individual cell(s).
- As used herein, to hybridize under conditions of a specified stringency is used to describe the stability of hybrids formed between two single-stranded DNA fragments and refers to the conditions of ionic strength and temperature at which such hybrids are washed, following annealing under conditions of stringency less than or equal to that of the washing step. Typically high, medium and low stringency encompass the following conditions or equivalent conditions thereto:
- 1) high stringency: 0.1×SSPE or SSC, 0.1% SDS, 65° C.
- 2) medium stringency: 0.2×SSPE or SSC, 0.1% SDS, 50° C.
- 3) low stringency: 1.0×SSPE or SSC, 0.1% SDS, 50° C. Equivalent conditions refer to conditions that select for substantially the same percentage of mismatch in the resulting hybrids. Additions of ingredients, such as formamide, Ficoll, and Denhardt's solution affect parameters such as the temperature under which the hybridization should be conducted and the rate of the reaction. Thus, hybridization in 5×SSC, in 20% formamide at 42° C. is substantially the same as the conditions recited above hybridization under conditions of low stringency. The recipes for SSPE, SSC and Denhardt's and the preparation of deionized formamide are described, for example, in Sambrook et al. (1989)Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Laboratory Press, Chapter 8; see, Sambrook et al, vol. 3, p. B.13, see, also, numerous catalogs that describe commonly used laboratory solutions). It is understood that equivalent stringencies can be achieved using alternative buffers, salts and temperatures.
- The terms substantially identical or similar varies with the context as understood by those skilled in the relevant art and generally means at least 40, 60, 80, 90, 95 or 98%.
- As used herein, substantially identical to a product means sufficiently similar so that the property of interest is sufficiently unchanged so that the substantially identical product can be used in place of the product.
- As used herein, target cell refers to a cell that expresses a cell surface protease.
- As used herein, test substance, including compounds provided herein, refers to a chemically defined compound (e.g., organic molecules, inorganic molecules, organic/inorganic molecules, proteins, peptides, nucleic acids, oligonucleotides, lipids, polysaccharides, saccharides, or hybrids among these molecules such as glycoproteins, etc.) or mixtures of compounds (e.g., a library of test compounds, natural extracts or culture supernatants, etc.) whose effect on or interaction with a cell surface protein or cell surface-associated protein, or a domain thereof, is determined by the methods herein.
- As used herein, the terms a therapeutic agent, therapeutic regimen, radioprotectant, chemotherapeutic mean conventional drugs and drug therapies, including vaccines, which are known to those skilled in the art. Radiotherapeutic agents are well known in the art.
- As used herein, vector (or plasmid) refers to discrete elements that are used to introduce heterologous DNA into cells for expression and/or replication thereof. The vectors typically remain episomal, but can be designed to effect integration of a gene or portion thereof into a chromosome of the genome. Also contemplated are vectors that are artificial chromosomes, such as yeast artificial chromosomes and mammalian artificial chromosomes. Selection and use of such vehicles are well known to those of skill in the art. An expression vector includes vectors capable of expressing DNA that is operatively linked with regulatory sequences, such as promoter regions, that are capable of effecting expression of such DNA fragments. Thus, an expression vector refers to a recombinant DNA or RNA construct, such as a plasmid, a phage, recombinant virus or other vector that, upon introduction into an appropriate host cell, results in expression of the cloned DNA. Appropriate expression vectors are well known to those of skill in the art and include those that are replicable in eukaryotic cells and/or prokaryotic cells and those that remain episomal or those which integrate into the host cell genome.
- As used herein, chemically stable means that the compound is stable enough to be formulated for pharmaceutical use. Such chemical stability is well known to those of skill in the art and can be determined by well known routine methods. Whether a given compound is chemically stable enough to be formulated for pharmaceutical use depends on a number of factors including, but not limited to, the type of formulation and route of administration desired, the disease to be treated, and the method of preparing the pharmaceutical formulation.
- As used herein, a “functional equivalent” of a side chain of an amino acid is a group or moiety that functions in substantially the same way as the naturally occurring side chain to achieve substantially the same result (e.g., a substrate for a cell surface protease). For example, functional equivalents of the side chain of arginine include, but are not limited to, homoarginine, guanidinoaminopropyl, guanidinoaminoethyl, (Me)2arginine side chain, (Et)2arginine side chain, (4-aminomethyl)phenylmethyl, 4-amidinophenylmethyl, 4-guanidinophenyl-methyl, or a conformationally constrained arginine side chain analog such as:
-
- where d is an integer from 0 to 5, or 1 to 3; and W is N or CH; or a mono- or di-substituted N-alkyl derivative of the above groups, where alkyl is, in certain embodiments, lower alkyl, such as methyl.
- As used herein, pharmaceutically acceptable derivatives of a compound include salts, esters, enol ethers, enol esters, acids, bases, solvates, hydrates or prodrugs thereof. Such derivatives can be readily prepared by those of skill in this art using known methods for such derivatization. The compounds produced can be administered to animals or humans without substantial toxic effects and either are pharmaceutically active or are prodrugs. Pharmaceutically acceptable salts include, but are not limited to, amine salts, such as but not limited to N,N′-dibenzylethylenediamine, chloroprocaine, choline, ammonia, diethanolamine and other hydroxyalkylamines, ethylenediamine, N-methylglucamine, procaine, N-benzylphenethylamine, 1-para-chlorobenzyl-2-pyrrolidin-1′-ylmethylbenzimidazole, diethylamine and other alkylamines, piperazine and tris(hydroxymethyl)aminomethane; alkali metal salts, such as but not limited to lithium, potassium and sodium; alkali earth metal salts, such as but not limited to barium, calcium and magnesium; transition metal salts, such as but not limited to zinc; and other metal salts, such as but not limited to sodium hydrogen phosphate and disodium phosphate; and also including, but not limited to, salts of mineral acids, such as but not limited to hydrochlorides and sulfates; and salts of organic acids, such as but not limited to acetates, lactates, malates, tartrates, citrates, ascorbates, succinates, butyrates, valerates and fumarates. Pharmaceutically acceptable esters include, but are not limited to, alkyl, alkenyl, alkynyl, aryl, heteroaryl, aralkyl, heteroaralkyl, cycloalkyl and heterocyclyl esters of acidic groups, including, but not limited to, carboxylic acids, phosphoric acids, phosphinic acids, sulfonic acids, sulfinic acids and boronic acids. Pharmaceutically acceptable enol ethers include, but are not limited to, derivatives of formula C═C(OR) where R is hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, aralkyl, heteroaralkyl, cycloalkyl or heterocyclyl. Pharmaceutically acceptable enol esters include, but are not limited to, derivatives of formula C═C(OC(O)R) where R is hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, aralkyl, heteroaralkyl, cycloalkyl ar heterocyclyl. Pharmaceutically acceptable solvates and hydrates are complexes of a compound with one or more solvent or water molecule, generally 1 to about 100, typically 1 to about 10, such as 1 to about 2, 3 or 4, solvent or water molecules.
- As used herein, treatment means any manner in which one or more of the symptoms of a condition, disorder or disease are ameliorated or otherwise beneficially altered. Treatment also encompasses any pharmaceutical use of the compositions herein, such as use for treating cancer.
- As used herein, amelioration of the symptoms of a particular disorder by administration of a particular pharmaceutical composition refers to any lessening, whether permanent or temporary, lasting or transient that can be attributed to or associated with administration of the composition.
- As used herein, a prodrug is a compound that, upon in vivo administration, is metabolized or otherwise converted to the biologically, pharmaceutically or therapeutically active form of the compound. To produce a prodrug, the pharmaceutically active compound is modified such that the active compound is regenerated by metabolic processes. The prodrug can be designed to alter the metabolic stability or the transport characteristics of a drug, to mask side effects or toxicity, to improve the flavor of a drug or to alter other characteristics or properties of a drug. By virtue of knowledge of pharmacodynamic processes and drug metabolism in vivo, those of skill in this art, once a pharmaceutically active compound is known, can design prodrugs of the compound (see, e.g., Nogrady (1985)Medicinal Chemistry A Biochemical Approach, Oxford University Press, New York, pages 388-392).
- It is to be understood that the conjugates provided herein can contain chiral centers. Such chiral centers can be of either the (R) or (S) configuration, or can be a mixture thereof. Thus, the compounds provided herein can be enantiomerically pure, or be stereoisomeric or diastereomeric mixtures. In the case of amino acid residues, such residues can be of either the L- or D-form. The configuration for naturally occurring amino acid residues is generally L. When not specified the residue is the L form. It is to be understood that the chiral centers of the compounds provided herein can undergo epimerization in vivo. As such, one of skill in the art will recognize that administration of a compound in its (R) form is equivalent, for compounds that undergo epimerization in vivo, to administration of the compound in its (S) form.
- The conjugates provided herein are prodrugs because they include a therapeutic agent in an inactive form that is ultimately converted to an active form at the targeted cell or tissue or in the environment thereof. Upon exposure to targeted protease either a biologically, pharmaceutically or therapeutically active form of a compound is released, or, a derivative that can be further metabolized into a biologically, pharmaceutically or therapeutically active form of a compound.
- As used herein, substantially pure means sufficiently homogeneous to appear free of readily detectable impurities as determined by standard methods of analysis, such as thin layer chromatography (TLC), gel electrophoresis, high performance liquid chromatography (HPLC) and mass spectrometry (MS), used by those of skill in the art to assess such purity, or sufficiently pure such that further purification would not alter the physical and chemical properties, such as enzymatic and biological activities, of the substance for its intended purpose. Methods for purification of the compounds to produce substantially chemically pure compounds are known to those of skill in the art. A substantially chemically pure compound may, however, be a mixture of stereoisomers. In such instances, further purification might increase the specific activity of the compound.
- As used herein, a peptidic substrate includes peptides and molecules, such as peptide mimetics and peptides that include peptide bond surrogates.
- As used herein, conventional terminology (Schecter et al. (1967)Biochem. Biophys. Res. Commun. 27:157-162) is used to refer to specific subsites of a protease substrate: Pn . . . P3-P2-P1↓P1′-P2′-P3′ . . . Pn′. The scissile bond (i.e., the cleavage site) of a substrate is indicated by the arrow. Positions N-terminal of that bond are referred to as unprimed positions. Subsites are then assigned a number based on their distance from the scissile bond. Amino acids (or amino acid surrogates) that form the scissile bond are assigned the
number 1, adjacent residues thenumber 2, and so on, counting away from the scissile bond. Each specific subsite of the substrate, therefore, is uniquely identified by a number and the designation as primed or unprimed. - As used herein, a surrogate of a peptide bond is a divalent group that possesses similar steric and/or electronic characteristics to —C(O)NH—. Peptide bond surrogates include, but are not limited to, alkene isosteres (—CR═CR—), particularly (E)-alkene isosteres of formula —CH═CH—, hydroxyethylene isosteres (—CH(OH)CH2—), enamine isosteres (—C(═CRR)NH—), aminoalcohol isosteres (—CH(OH)CH2NH—), difluoroketone isosteres (—C(O)CF2—), retroinverso compounds (—NHC(O)—), divalent heterocyclyl or heteroaryl groups, and cyclopropyl isosteres such as:
- As used herein, alkyl, alkenyl and alkynyl carbon chains, if not specified, contain from 1 to 20 carbons, generally 1 to 16 carbons, and are straight or branched. Alkenyl carbon chains of from 2 to 20 carbons typically contain 1 to 8 double bonds, and the alkenyl carbon chains of 2 to 16 carbons and typically contain 1 to 5 double bonds. Alkynyl carbon chains of from 2 to 20 carbons typically contain 1 to 8 triple bonds, and the alkynyl carbon chains of 2 to 16 carbons and generally contain 1 to 5 triple bonds. Exemplary alkyl, alkenyl and alkynyl groups herein include, but are not limited to, methyl, ethyl, propyl, isopropyl, isobutyl, n-butyl, sec-butyl, tert-butyl, isopentyl, neopentyl, tert-pentyl and isohexyl. The alkyl, alkenyl and alkynyl groups, unless otherwise specified, optionally can be substituted, with one or more groups, generally alkyl group substituents that are the same or different. As used herein, lower alkyl, lower alkenyl, and lower alkynyl refer to carbon chains having less than about 6 carbons. As used herein, “alk(en)(yn)yl” refers to an alkyl group containing at least one double bond and at least one triple bond.
- As used herein, “cycloalkyl” refers to a saturated mono- or multi-cyclic ring system, typically 3 to 10 carbon atoms, such as, for example, 3 to 6 carbon atoms; cycloalkenyl and cycloalkynyl refer to mono- or multicyclic ring systems that respectively include at least one double bond and at least one triple bond. Cycloalkenyl and cycloalkynyl groups contain, for example, 3 to 10 carbon atoms, with cycloalkenyl groups generally containing 4 to 7 carbon atoms and cycloalkynyl groups that contain, for example 8 to 10 carbon atoms. The ring systems of the cycloalkyl, cycloalkenyl and cycloalkynyl groups can be composed of one ring or two or more rings which can be joined together in a fused, bridged or spiro-connected fashion, and optionally can be substituted with one or more alkyl group substituents. “Cycloalk(en)(yn)yl” refers to a cycloalkyl group containing at least one double bond and at least one triple bond.
- As used herein, “substituted alkyl,” “substituted alkenyl,” “substituted alkynyl,” “substituted cycloalkyl,” “substituted cycloalkenyl,” and “substituted cycloalkynyl” refer to alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl and cycloalkynyl groups, respectively, that are substituted with one or more substituents, in certain embodiments one to three substituents, independently selected from alkyl, halo, haloalkyl, such as halo lower alkyl, pseudohalo, aryl, amino, dialkylamino, nitro, cyano, azido, alkylsulfinyl, alkylsulfonyl, alkylcarbonylamino, alkoxycarbonylamino, aminoimino, hydroxy, alkoxy, aryloxy, alkyloxy, alkylthio, arylthio, aralkyloxy, aralkylthio, carboxy, alkylcarbonyl, alkoxycarbonyl, oxo and cycloalkyl.
- As used herein, “aryl” refers to cyclic groups containing from 6 to 19 carbon atoms. Aryl groups include, but are not limited to groups, such as fluorenyl, substituted fluorenyl, phenyl, substituted phenyl, naphthyl and substituted naphthyl. As used herein, “aryl” also refers to aryl-containing groups, including, but not limited to, aryloxy, arylthio, arylcarbonyl and arylamino groups.
- As used herein, “heteroaryl” refers to a monocyclic or multicyclic aromatic ring system, generally about 5 to about 15 members where one or more, such as 1 to 3 of the atoms in the ring system is a heteroatom, that is, an element other than carbon, for example, nitrogen, oxygen and sulfur atoms. The heteroaryl group optionally can be fused to a benzene ring. Exemplary heteroaryl groups include, for example, furyl, imidazolyl, pyrrolidinyl, pyrimidinyl, tetrazolyl, thienyl, pyridyl, pyrrolyl, N-methylpyrrolyl, quinolinyl and isoquinolinyl, with pyridyl, thienyl and quinolinyl as examples thereof.
- As used herein, “heteroaryl” also refers to heteroaryl-containing groups, including, but not limited to, heteroaryloxy, heteroarylthio, heteroarylcarbonyl and heteroarylamino.
- As used herein, “heterocyclyl” refers to a monocyclic or multicyclic non-aromatic ring system, such as systems of 3 to 10 members, for
exmaple 4 to 7 members or 5 to 6 members, where one or more, such as 1 to 3 of the atoms in the ring system is a heteroatom, that is, an element other than carbon, for example, nitrogen, oxygen and/or sulfur atoms. - As used herein, “substituted aryl,” “substituted heteroaryl” and “substituted heterocyclyl” refer to aryl, heteroaryl and heterocyclyl groups, respectively, that are substituted with one or more substituents, in certain embodiments one to three substituents, independently selected from alkyl, cycloalkyl, cycloalkylalkyl, aryl, heteroaryl optionally substituted with 1 or more, such as 1 to 3, substituents selected from halo, halo alkyl and alkyl, aralkyl, heteroaralkyl, alkenyl containing 1 to 2 double bonds, alkynyl containing 1 to 2 triple bonds, alk(en)(yn)yl groups, halo, pseudohalo, cyano, hydroxy, haloalkyl and polyhaloalkyl, such as halo lower alkyl, especially trifluoromethyl, formyl, alkylcarbonyl, arylcarbonyl that optionally is substituted with 1 or more, generally 1 to 3, substituents selected from halo, halo alkyl and alkyl, heteroarylcarbonyl, carboxy, alkoxycarbonyl, aryloxycarbonyl, aminoimino, alkoxycarbonylamino, aryloxycarbonylamino, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, arylaminocarbonyl, diarylaminocarbonyl, aralkylaminocarbonyl, alkoxy, aryloxy, perfluoroalkoxy, alkenyloxy, alkynyloxy, arylalkoxy, aminoalkyl, alkyl-aminoalkyl, dialkylaminoalkyl, arylaminoalkyl, amino, alkylamino, dialkyl-amino, arylamino, alkylarylamino, alkylcarbonylamino, arylcarbonylamino, azido, nitro, mercapto, alkylthio, arylthio, perfluoroalkylthio, thiocyano, isothiocyano, alkylsulfinyl, alkylsulfonyl, arylsulfinyl, arylsulfonyl, aminosulfonyl, alkylaminosulfonyl, dialkylaminosulfonyl and arylamino-sulfonyl.
- As used herein, “aralkyl” refers to an alkyl group in which one of the hydrogen atoms of the alkyl is replaced by an aryl group.
- As used herein, “heteroaralkyl” refers to an alkyl group in which one of the hydrogen atoms of the alkyl is replaced by a heteroaryl group.
- As used herein, the nomenclature alkyl, alkoxy, carbonyl, etc. is used as is generally understood by those of skill in this art. For example, as used herein alkyl refers to saturated carbon chains that contain one or more carbons; the chains can be straight or branched or include cyclic portions or be cyclic.
- Where the number of any given substituent is not specified (e.g., “haloalkyl”), there can be one or more substituents present. For example, “haloalkyl” can include one or more of the same or different halogens. As another example, “C1-3alkoxyphenyl” can include one or more of the same or different alkoxy groups containing one, two or three carbons.
- As used herein, “halo”, “halogen” or “halide” refers to F, Cl, Br or I.
- As used herein, pseudohalides are compounds that behave substantially similar to halides. Such compounds can be used in the same manner and treated in the same manner as halides (X−, in which X is a halogen, such as Cl or Br). Pseudohalides include, but are not limited to, cyanide, cyanate, thiocyanate, selenocyanate, trifluoromethoxy, difluoromethoxy, dichloromethoxy and azide.
- As used herein, “haloalkyl” refers to a lower alkyl radical in which one or more of the hydrogen atoms are replaced by halogen. Such groups include, but not limited to, chloromethyl, trifluoromethyl, 1-chloro-2-fluoroethyl and the like.
- As used herein, “haloalkoxy” refers to RO— in which R is a haloalkyl group.
- As used herein, “sulfinyl” or “thionyl” refers to —S(O)—. As used herein, “sulfonyl” or “sulfuryl” refers to —S(O)2—. As used herein, “sulfo” refers to —S(O)2O—.
- As used herein, “carboxy” refers to a divalent radical, —C(O)O—.
- As used herein, “aminocarbonyl” refers to —C(O)NH2.
- As used herein, “alkylaminocarbonyl” refers to —C(O)NHR in which R is hydrogen or alkyl, such as, for example, lower alkyl.
- As used herein “dialkylaminocarbonyl” as used herein refers to —C(O)NR′R in which R′ and R are independently selected from hydrogen or alkyl, such as, for example, lower alkyl; “carboxamide” refers to groups of formula —NR′COR.
- As used herein, “diarylaminocarbonyl” refers to —C(O)NRR′ in which R and R′ are independently selected from aryl, such as lower aryl, for example, phenyl.
- As used herein, “aralkylaminocarbonyl” refers to —C(O)NRR′ in which one of R and R′ is aryl, such as, lower aryl, for example, phenyl, and the other of R and R′ is alkyl, such as, for example, lower alkyl.
- As used herein, “arylaminocarbonyl” refers to —C(O)NHR in which R is aryl, such as lower aryl, for example, phenyl.
- As used herein, “hydroxycarbonyl” refers to —COOH.
- As used herein, “alkoxycarbonyl” refers to —C(O)OR in which R is alkyl, such as lower alkyl.
- As used herein, “aryloxycarbonyl” refers to —C(O)OR in which R is aryl, such lower aryl, for example phenyl.
- As used herein, “alkoxy” and “alkylthio” refer to RO— and RS—, in which R is alkyl, such as, for example, lower alkyl.
- As used herein, “aryloxy” and “arylthio” refer to RO— and RS—, in which R is aryl, such lower aryl, for example, phenyl.
- As used herein, “alkylene” refers to a straight, branched or cyclic, such as, for example, straight or branched, divalent aliphatic hydrocarbon group, for example, having from 1 to about 20 carbon atoms such as 1 to 12 carbons, and for exmaple, is lower alkylene. There optionally can be inserted along the alkylene group one or more oxygen, sulphur or substituted or unsubstituted nitrogen atoms, where the nitrogen substituent is alkyl as previously described. Exemplary alkylene groups include methylene (—CH2—), ethylene (—CH2CH2—), propylene (—(CH2)3—), cyclohexylene (—C6H10—), methylenedioxy (—O—CH2—O—) and ethylenedioxy (—O—(CH2)2—O—). The term “lower alkylene” refers to alkylene groups having 1 to 6 carbons. Exemplary alkylene groups are lower alkylene, such as, for example, alkylene of 1 to 3 carbon atoms.
- As used herein, “alkenylene” refers to a straight, branched or cyclic, typically straight or branched, divalent aliphatic hydrocarbon group, such as, for example, having from 2 to about 20 carbon atoms and at least one double bond, generally 1 to 12 carbons, and is for example, lower alkenylene. There optionally can be inserted along the alkenylene group one or more oxygen, sulphur or substituted or unsubstituted nitrogen atoms, where the nitrogen substituent is alkyl as previously described. Exemplary alkenylene groups include —CH═CH—CH═CH— and —CH═CH═CH2—. The term “lower alkenylene” refers to alkenylene groups having 2 to 6 carbons. Examplary alkenylene groups are lower alkenylene, such as, for example, alkenylene of 3 to 4 carbon atoms.
- As used herein, “alkynylene” refers to a straight, branched or cyclic, generally straight or branched, divalent aliphatic hydrocarbon group, such those having from 2 to about 20 carbon atoms and at least one triple bond, generally 1 to 12 carbons, such as, for example, lower alkynylene. There optionally can be inserted along the alkynylene group one or more oxygen, sulphur or substituted or unsubstituted nitrogen atoms, where the nitrogen substituent is alkyl as previously described. Exemplary alkynylene groups include —C≡C—C≡C—, —C≡C— and —C≡C—CH2—. The term “lower alkynylene” refers to alkynylene groups having 2 to 6 carbons. Exemplary alkynylene groups are lower alkynylene, such as, for example, alkynylene of 3 to 4 carbon atoms.
- As used herein, “alk(en)(yn)ylene” refers to a straight, branched or cyclic, generally straight or branched, divalent aliphatic hydrocarbon group, having, for example, from 2 to about 20 carbon atoms and at least one triple bond, and at least one double bond; typically 1 to 12 carbons, such as, for example, lower alk(en)(yn)ylene. There optionally can be inserted along the alkynylene group one or more oxygen, sulphur or substituted or unsubstituted nitrogen atoms, where the nitrogen substituent is alkyl as previously described. Exemplary alk(en)(yn)ylene groups include —C═C—(CH2)n—C≡C—, where n is 1 or 2. The term “lower alk(en)(yn)ylene” refers to alk(en)(yn)ylene groups having up to 6 carbons. Exemplary alk(en)(yn)ylene groups are lower alk(en)(yn)ylene, such as, for example, alk(en)(yn)ylene of 4 carbon atoms.
- As used herein, “cycloalkylene” refers to a divalent saturated mono- or multicyclic ring system, generally 3 to 10 carbon atoms, such as 3 to 6 carbon atoms; cycloalkenylene and cycloalkynylene refer to divalent mono- or multicyclic ring systems that respectively include at least one double bond and at least one triple bond. Cycloalkenylene and cycloalkynylene groups can contain 3 to 10 carbon atoms, with, for example, cycloalkenylene groups containing 4 to 7 carbon atoms and cycloalkynylene groups containing 8 to 10 carbon atoms. The ring systems of the cycloalkylene, cycloalkenylene and cycloalkynylene groups can be composed of one ring or two or more rings that can be joined together in a fused, bridged or spiro-connected fashion. “Cycloalk(en)(yn)ylene” refers to a cycloalkylene group containing at least one double bond and at least one triple bond.
- As used herein, “substituted alkylene,” “substituted alkenylene,” “substituted alkynylene,” “substituted cycloalkylene,” “substituted cycloalkenylene,” and “substitued cycloalkynylene” refer to alkylene, alkenylene, alkynylene, cycloalkylene, cycloalkenylene and cycloalkynylene groups, respectively, that are substituted with one or more substituents, in certain embodiments one to three substituents, independently selected from halo, haloalkyl, such as, for example, halo lower alkyl, aryl, hydroxy, alkoxy, aryloxy, alkyloxy, alkylthio, arylthio, aralkyloxy, aralkylthio, carboxy alkoxycarbonyl, oxo and cycloalkyl.
- As used herein, “arylene” refers to a monocyclic or polycyclic, such as monocyclic, divalent aromatic group, for example, having from 5 to about 20 carbon atoms and at least one aromatic ring, such as 5 to 12 carbons, and, is, for example, lower arylene. There optionally can be inserted around the arylene group one or more oxygen, sulphur or substituted or unsubstituted nitrogen atoms, where the nitrogen substituent is alkyl as previously described. Exemplary arylene groups include 1,2-, 1,3- and 1,4-phenylene. The term “lower arylene” refers to arylene groups having 5 or 6 carbons. Exemplary arylene groups are lower arylene.
- As used herein, “heteroarylene” refers to a divalent monocyclic or multicyclic aromatic ring system, such as of about 5 to about 15 members where one or more, typically, for example, 1 to 3 of the atoms in the ring system is a heteroatom, that is, an element other than carbon, for example, nitrogen, oxygen and/or sulfur atom(s).
- As used herein, “heterocyclylene” refers to a divalent monocyclic or multicyclic non-aromatic ring system, generally of 3 to 10 members, such as, for example, 4 to 7 members or 5 to 6 members, where one or more, such as, for example, 1 to 3 of the atoms in the ring system is a heteroatom, that is, an element other than carbon, for example, nitrogen, oxygen and/or sulfur atom(s).
- As used herein, “substituted arylene,” “substituted heteroarylene” and “substituted heterocyclylene” refer to arylene, heteroarylene and heterocyclylene groups, respectively, that are substituted with one or more substituents, in certain embodiments one to three substituents, independently selected from alkyl, cycloalkyl, cycloalkylalkyl, aryl, heteroaryl optionally substituted with 1 or more, such as 1 to 3, substituents selected from halo, halo alkyl and alkyl, aralkyl, heteroaralkyl, alkenyl containing 1 to 2 double bonds, alkynyl containing 1 to 2 triple bonds, alk(en)(yn)yl groups, halo, pseudohalo, cyano, hydroxy, haloalkyl and polyhaloalkyl, such as, halo lower alkyl, for example trifluoromethyl, formyl, alkylcarbonyl, arylcarbonyl that optionally is substituted with 1 or more, such as 1 to 3, substituents selected from, for example, halo, halo alkyl and alkyl, heteroarylcarbonyl, carboxy, alkoxycarbonyl, aryloxycarbonyl, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, arylaminocarbonyl, diarylaminocarbonyl, aralkylaminocarbonyl, alkoxy, aryloxy, perfluoroalkoxy, alkenyloxy, alkynyloxy, arylalkoxy, aminoalkyl, alkylaminoalkyl, dialkylaminoalkyl, arylaminoalkyl, amino, alkylamino, dialkylamino, arylamino, alkylarylamino, alkylcarbonylamino, arylcarbonylamino, azido, nitro, mercapto, alkylthio, arylthio, perfluoroalkylthio, thiocyano, isothiocyano, alkylsulfinyl, alkylsulfonyl, arylsulfinyl, arylsulfonyl, aminosulfonyl, alkylaminosulfonyl, dialkylaminosulfonyl and arylaminosulfonyl.
- As used herein, “alkylidene” refers to a divalent group, such as ═CR′R″, which is attached to one atom of another group, forming a double bond. Exemplary alkylidene groups are methylidene (═CH2) and ethylidene (═CHCH3). As used herein, “aralkylidene” refers to an alkylidene group in which either R′ or R″ is an aryl group. “Cycloalkylidene” groups are those where R′ and R″ are linked to form a carbocyclic ring. “Heterocyclylidene” groups are those where at least one of R′ and R″ contain a heteroatom in the chain, and R′ and R″ are linked to form a heterocyclic ring.
- As used herein, “amido” refers to the divalent group —C(O)NH—. “Thioamido” refers to the divalent group —C(S)NH—. “Oxyamido” refers to the divalent group —OC(O)NH—. “Thiaamido” refers to the divalent group —SC(O)NH—. “Dithiaamido” refers to the divalent group —SC(S)NH—. “Ureido” refers to the divalent group —HNC(O)NH—. “Thioureido” refers to the divalent group —HNC(S)NH—.
- As used herein, “semicarbazide” refers to —NHC(O)NHNH—. “Carbazate” refers to the divalent group —OC(O)NHNH—. “Isothiocarbazate” refers to the divalent group —SC(O)NHNH—. “Thiocarbazate” refers to the divalent group —OC(S)NHNH—. “Sulfonylhydrazide” refers to the group —SO2NHNH—. “Hydrazide” refers to the divalent group —C(O)NHNH—. “Azo” refers to the divalent group —N═N—. “Hydrazinyl” refers to the divalent group —NH—NH—.
- As used herein, the term “amino acid” refers to α-amino acids which are racemic, or of either the D- or L-configuration. The designation “d” preceding an amino acid designation (e.g., dAla, dSer, dVal, etc.) refers to the D-isomer of the amino acid. The designation “dI” preceding an amino acid designation (e.g., dIPip) refers to a mixture of the L- and D-isomers of the amino acid.
- As used herein, when any particular group, such as phenyl or pyridyl, is specified, this means that the group is unsubstituted or is substituted. Exemplary substituents where not specified are halo, halo lower alkyl, and lower alkyl.
- As used herein, the abbreviations for any protective groups, amino acids and other compounds, are, unless indicated otherwise, in accord with their common usage, recognized abbreviations, or the IUPAC-IUB Commission on Biochemical Nomenclature (see, (1972)Biochem. 11:942-944).
- As used herein, HHT and CHT refer to hexahydrotyrosyl (also known as cyclohexyltyrosyl or p-hydroxycyclohexylalanyl), CHA is cyclohexylalanyl, Pyr and pyroGlu refer to pyroglutamic acid, Pip is pipecolinic acid, Sar is sarcosine, nLeu and Nle are norleucine, nVal is norvaline, Aib is 2-aminoisobutyric acid, Quat is (R)-Glu(α-(3-amidinobenzyl)), and Abu and But are 2-aminobutyric acid.
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- As used herein, the term “hydroxylated” represents substitution on a substitutable carbon of the ring system being so described by a hydroxyl moiety. As used herein, the term “polyhydroxylated” represents substitution on two or more substitutable carbons of the ring system being so described by 2, 3 or 4 hydroxyl moieties.
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- or a diastereomer thereof.
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- or a diastereomer thereof.
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- or a diastereomer thereof.
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- and
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- Other abbreviations as used herein are as follows:
Abbreviation Refers to Aib 2- aminoisobutyryl 4,4-dimethylThr 2-amino-3-hydroxy-4-methylpentanoyl Met(O2) methioninyl-S,S-dioxide Ser(OMe) the O-methyl ether of serinyl, also known as 2-amino-3-methoxypropanoyl hSer homoserinyl, also known as 2-amino-4- hydroxybutanoyl (hS)Gly N-(2-hydroxyethyl)glycyl N,N-dimethylGly N,N-dimethylglycyl β-Ala 3-aminopropanoyl Cys(Me) S-methylcysteinyl t-butylGly 2-amino-3,3-dimethylbutanoyl F(Gn) 4-guanidinyiphenylalanyl hCHA homocyclohexylalanyl, or 2-amino-4- cyclohexylbutanoyl hexylGly 2-aminooctanoyl allylGly 2-amino-4-pentenoyl Inact. inactive NT not tested MeOEtCO 3- methoxypropanoyl 3,4- MethyldioxyPhAc 3,4-methylenedioxyphenylacetyl L-3-PhLactyl L-2-hydroxy-3-phenylpropanoyl MeOEtOCO 2-methoxyethoxycarbonyl MeOCO methoxycarbonyl MeO(EtO)2Ac 2-(2-methoxyethoxy)ethoxyacetyl 2-PyridylAc 2-pyridylacetyl PhOAc phenoxyacetyl MeOAc methoxyacetyl PhAc phenylacetyl MeOEtOAc 2-methoxyethoxyacetyl HOOCButa glutaryl Z benzyloxycarbonyl EtOCO ethoxycarbonyl βA beta-alanyl or 3-aminopropanoyl NapAc 1-naphthylacetyl iBoc isobutoxycarbonyl HOAc hydroxyacetyl MeSucc 3-methoxycarbonylpropanoyl Succ succinyl HCO formyl 4-(guan)Phg 4-guanidinylphenylglycyl Dox doxorubicin Tax taxol dA(Chx) or dCha d-cyclohexylalanyl dhF d-homophenylalanyl P(OH) 4-hydroxyprolyl - The conjugates herein are designed to target proteases that are located on cell surfaces, particularly tumor cells and cells involved in tumorigenic processes and angiogenesis and other proliferative processes. The conjugates, described in detail below, contain a peptidic substrate for a selected targeted cell surface protease linked, either directly or via a linker, to a therapeutic agent, typically a cytotoxic agent, which is substantially inactive when in the conjugate. The therapeutic agent is released in a form that is active or that can be activated in the vicinity of the targeted cell or tissue to which it is delivered. As a result, active therapeutic agent accumulates at the targeted cells or tissue or in the targeted cells.
- The targeted protease is selected by identifying a protease that is located on a cell or tissue (or associated therewith) that is involved in the disease process or serendipitously present in the locale of cells or tissues involved in the disease or disease process, and, generally, is not located at all or present or active at lower levels, generally substantially lower levels, or exhibits altered activity or specificity, on many, if not all, other cells or tissues. The variety and numbers of non-targeted cells or tissues that expresss the active protease varies for particular proteases and diseases intended for treatment. Those of skill in the art will select a target based upon the disease, targeted agents and tolerable or acceptable levels of side-effects. The goal is to achieve enhanced therapeutic index compared with administration of the targeted agent by itself.
- The targeted protease may or may not be involved in the disease process and its expression can be serendiptous; for purposes herein its particular role or lack thereof is not important; it is the fact that it is active in the locale of targeted tissues or cells that is important. For example, many of the cell surface proteases of interest herein are expressed or active on tumor cells or cells involved in the tumorigenic processes. Any method known to one of skill in the art for determining or detecting a tissue or cell expression profile can be used. For example, RNA blots composed of RNA from numerous tissues (e.g., a multiple tissue expression (MTE) array available from CLONTECH, Palo Alto, Calif.), can be screened with probes based upon the nucleic acid sequence of the protease of interest to identify cells that express the protease. Northern analysis of the blots to test for expression also can be used.
- Included among the targeted proteases are those designated type II membrane-bound serine proteases (MTSPs; see, e.g., U.S. application Ser. No. 09/776,191, filed Feb. 2, 2001 and International PCT application No. PCT/US01/03471 published as International PCT application No. WO 01/57194; see International PCT application No. PCT/US02/07903; see, also U.S. provisional application Serial Nos. 60/275,592, 60/278,166, 60/279,228, 60/291,001, 60/291,501 60/316,818, 60/302,939, 60/316,818, 60/328,529, 60/328,530, 60/332,015, 60/328,939, and provisional application, filed on May 20, 2002 under attorney docket no. 24745-P1624; U.S. application Ser. Nos. 10/099,700, 10/104,271, 10/112,221, application filed on May 14, 2002 under attorney docket no. 24745-1616) and those found on endothelial cells designated endotheliases (see, U.S. application Ser. No. 09/717,473, filed Nov. 20, 2000, and International PCT application No. PCT/US00/31803 published as International PCT application No. WO 01/36604); see, also SEQ ID Nos. 3-26, 269-270 and 272-276.
- Also contemplated are proteases that are located at the cell surface by virtue of a specific interaction with a cell surface protein. Urokinase plasminogen activator (u-PA) bound to urokinase plasminogen activator receptor (u-PAR) is exemplary of such proteases. Nucleic acid sequence information and expression profiles of exemplary MTSPs and endotheliases are as follows (see, also EXAMPLE 6).
- 1. MTSPs
- Cell surface proteolysis is a mechanism for the generation of biologically active proteins that mediate a variety of cellular functions. These membrane-anchored proteins, include a disintegrin-like and metalloproteinase (ADAM) and membrane-type matrix metalloproteinase (MT-MMP). In addition to the MMPs, serine proteases have been implicated in neoplastic disease progression. Most serine proteases, which are either secreted enzymes or are sequestered in cytoplasmic storage organelles, have roles in blood coagulation, wound healing, digestion, immune responses and tumor invasion and metastasis.
- Transmembrane serine proteases (MTSPs) appear to be involved in the etiology and pathogenesis of tumors. These enzymes are expressed in certain cancerous and tumor cells and in other cells associated with other proliferative disorders and other disease states, such as in inflammatory cells and and can be tissue or organ-specific. In mammals, more than 20 members of the family are known (see, Hooper et al. (2001)J. Biol. Chem. 276:857-860, see, also U.S. application Ser. No. 09/776,191, filed Feb. 2, 2001 and International PCT application No. PCT/US01/03471; see, also U.S. provisional application Serial Nos. 60/275,592 and 60/278,166; and see SEQ ID Nos. 1-37). These include corin (accession nos. AF133845 and AB013874; see, Yan et al. (1999) J. Biol. Chem. 274:14926-14938; Tomia et al. (1998) J. Biochem. 124:784-789; Uan et al. (2000) Proc. Natl. Acad. Sci. U.S.A. 97:8525-8529); enterpeptidase (also designated enterokinase; accession no. U09860 for the human protein; see, Kitamoto et al. (1995) Biochem. 27: 4562-4568; Yahagi et al. (1996) Biochem. Biophys. Res. Commun. 219:806-812; Kitamoto et al. (1994) Proc. Natl. Acad. Sci. U.S.A. 91:7588-7592; Matsushima et al. (1994) J. Biol. Chem. 269:19976-19982;); human airway trypsin-like protease (HAT; accession no. AB002134; see Yamaoka et al. J. Biol. Chem. 273:11894-11901); MTSP1 (also called TADG-15 and matriptase, see SEQ ID Nos. 1 and 2; accession nos. AF133086/AF118224, AF04280022; Takeuchi et al. (1999) Proc. Natl. Acad. Sci. U.S.A. 96:11054-1161; Lin et al. (1999) J. Biol. Chem. 274:18231-18236; Takeuchi et al. (2000) J. Biol. Chem. 275:26333-26342; and Kim et al. (1999) Immunogenetics 49:420-429); hepsin (see, accession nos. M18930, AF030065, X70900; Leytus et al. (1988) Biochem. 27: 11895-11901; Vu et al. (1997) J. Biol. Chem. 272:31315-31320; and Farley et al. (1993) Biochem. Biophys. Acta 1173:350-352; and see, U.S. Pat. No. 5,972,616); TMPRS2 (see, Accession Nos. U75329 and AF113596; Paoloni-Giacobino et al. (1997) Genomics 44:309-320; and Jacquinet et al. (2000) FEBS Lett. 468: 93-100); and TMPRSS4 (see, Accession No. NM 016425; Wallrapp et al. (2000) Cancer 60:2602-2606). Also known MTSP3, MTSP4, MTSP6, MTSP7, MTSP9, MTSP10, MTSP12, MTSP20, MTSP22 and MTSP25 (see, SEQ ID NOs. 3-26, 269-270 and 272-276; see, also U.S. application Ser. No. 09/776,191, filed Feb. 2, 2001 and International PCT application No. PCT/US01/03471 published as International PCT application No. WO 01/57194; see International PCT application No. PCT/US02/07903; see, also U.S. provisional application Serial Nos. 60/275,592, 60/278,166, 60/279,228, 60/291,001, 60/291,50160/316,818, 60/302,939, 60/316,818, 60/328,529, 60/328,530, 60/332,015, 60/328,939, and provisional application, filed on May 20 2002, under attorney docket no. 24745-P1624; U.S. application Ser. Nos. 10/099,700, 10/104,271, 10/112,221, application filed on May 14, 2002 under attorney docket no. 24745-1616)).
- Serine proteases, including transmembrane serine proteases, have been implicated in processes involved in neoplastic development and progression. While the precise role of these proteases has not been elaborated, serine proteases and inhibitors thereof are involved in the control of many intra- and extracellular physiological processes, including degradative actions in cancer cell invasion, metastatic spread, and neovascularization of tumors, that are involved in tumor progression. It is believed that proteases are involved in the degradation of extracellular matrix (ECM) and contribute to tissue remodeling, and are necessary for cancer invasion and metastasis. The activity and/or expression of some proteases have been shown to correlate with tumor progression and development, and also are shown to be active in specific cell types.
- For example, a membrane-type serine protease MTSP1 (also called matriptase; see SEQ ID Nos. 1 and 2 from U.S. Pat. No. 5,972,616; and GenBank Accession No. AF118224; (1999)J. Biol. Chem. 274:18231-18236; U.S. Pat. No. 5,792,616; see, also Takeuchi (1999) Proc. Natl. Acad. Sci. U.S.A. 96:11054-1161) that is expressed in epithelial cancer and normal tissue (Takeucuhi et al. (1999) Proc. Natl. Acad. Sci. USA 96:11054-61) has been identified. It has been proposed that it plays a role in the metastasis of breast cancer. Its primary cleavage specificity is Arg-Lys residues. Matriptase also is expressed in a variety of epithelial tissues with high levels of activity and/or expression in the human gastrointestinal tract and the prostate.
- Hepsin, a cell surface serine protease identified in hepatoma cells, is overexpressed in ovarian cancer (Tanimoto et al. (1997)Cancer Res., 57:2884-7). The hepsin transcript appears to be abundant in carcinoma tissue and is almost never expressed in normal adult tissue, including normal ovary. It has been suggested that hepsin is frequently overexpressed in ovarian tumors and therefore can be a candidate protease in the invasive process and growth capacity of ovarian tumor cells.
- A serine protease-like gene, designated normal epithelial cell-specific 1 (NES1) (Liu et al. (1996)Cancer Res. 56:3371-9) has been identified. Although expression of the NES1 mRNA is observed in all normal and immortalized nontumorigenic epithelial cell lines, the majority of human breast cancer cell lines show a drastic reduction or a complete lack of its expression. The structural similarity of NES1 to polypeptides known to regulate growth factor activity and a negative correlation of NES1 expression with breast oncogenesis suggest a direct or indirect role for this protease-like gene product in the suppression of tumorigenesis.
- Exemplary MTSPs
- Each MTSP has a characteristic tissue expression profile; the MTSPs in particular, although not exclusively expressed or activated in tumors, exhibit characteristic tumor tissue expression or activation profiles. In some instances, MTSPs can have different activity in a tumor cell from a non-tumor cell by virtue of a change in a substrate or cofactor therefor or other factor that would alter functional activity of the MTSP. Hence each can serve as a diagnostic marker for particular tumors, by virtue of a level of activity and/or expression or function in a subject (i.e. a mammal, particularly a human) with neoplastic disease, compared to a subject or subjects that do not have the neoplastic disease. In addition, detection of activity (and/or expression) in a particular tissue can be indicative of neoplastic disease. Also, by virtue of the activity and/or expression profiles of each, they can serve as therapeutic targets, such as by administration of modulators of the activity thereof, or, as by administration of a prodrug specifically activated by one of the MTSPs. Each or any of the MTSPs can exhibit activity or expression levels or substrate specificities that differ in tumor cells from the levels in normal cells. Such tumor cells include, but are not limited to, colon, lung, prostate, breast, esophagous, pancreas, cervic, uterus, endometrium, and other solid tumors and in blood and lymphatic tumors. Hence, conjugates provided herein can be designed by selection of substrate specificity for treatment of any of such tumors and neoplastic conditions.
- Tissue Expression Profiles
- The following are exemplary tissue and gene (see also, EXAMPLE 8) profiles of some exemplary MTSPs. These profiles are not intended to define the full scope of expression or activation of these MTPSs, but demonstrate that MTSPs are expressed in tumors, and, hence there expression or activation or substrate specificity on the surface of tumor cells can be exploited in the methods herein and conjugates, designed in accord with the methods herein and as exemplified herein, that are cleaved by one or more of these MTSPs can be prepared and employed for treatment of neoplastic or other diseases or conditions or to target to cells that express these proteins on there surfaces.
- MTSP1 (Matriptase)
- MTSP1 (also called matriptase) is a trypsin-like serine protease with broad spectrum cleavage activity and two potential regulatory modules. It was named “matriptase” based on its ability to degrade the extra-cellular matrix and its trypsin-like activity. When isolated from breast cancer cells (or T-47D cell conditioned medium), MTSP1 has been reported to be primarily in an uncomplexed form. MTSP1 has been isolated from human milk; when isolated from human milk, it was reported to be in one of two complexed forms, 95 kDa (the predominant form) and 110 kDa; uncomplexed MTSP1 was not detected (Liu, et al. (1999)J. Biol. Chem. 274:18237-18242). It has been proposed that MTSP1 exists as an uncomplexed protease when in its active state. In breast milk, it has been reported to exist in complex with a fragment of hepatocyte growth factor inhibitor-1 (HAI-1), a Kunitz-type serine protease inhibitor having activity against trypsin-like serine proteases.
- Nucleic acids encoding the protein designed matriptase were cloned from T-47D human breast cancer cell-conditioned medium (Lin et al. (1999)J. Biol. Chem. 274:18231-18236). Upon analysis of the cDNA, it was determined that the full length protease has 683 amino acids and contains three main structural regions: a serine protease domain near the carboxyl-terminal region, four tandem low-density lipoprotein receptor domains, and two tandem complement subcomponents C1r and C1s (see SEQ ID No. 1). Studies to identify additional serine proteases made by cancer cells were done using PC-3 cells. A serine protease termed “MT-SP1” (MTSP1) by the authors, reported to be a transmembrane protease was cloned (Takeuchi et al. (1999) Proc. Natl. Acad. Sci. U.S.A. 96:11054-11061). It was subsequently found that originally identified matriptase sequence is included in the translated sequence of the cDNA that encodes MTSP1. The nucleic acid encoding the protein originally designated matriptase is a partial MTSP1 clone that lacks 516 of the coding nucleotides (Takeuchi, et al., J. Biol. Chem 275:26333-26342 (2000).) Since the reported matriptase encoding cDNA sequence encoded a possible initiating methionine, it was proposed that alternative splicing could yield a protein lacking the N-terminal region of MTSP1. Hence, matriptase herein is a variant form of MTSP1.
- MTSP1 demonstrates trypsin-like protease activity and is a Type II transmembrane protein with an extracellular protease domain. Studies of substrate specificity of MTSP1 reveal that protease-activated receptor 2 (PAR2), pro-hepatacyte growth factor (pro-HGF) and single-chain urokinase-type plasminogen activator (sc-uPA) are macromolecular substrates of MTSP1. PAR2 functions in inflammation, cytoprotection and/or cell adhesion, while sc-uPa functions in tumor cell invasion and metastasis. HGF serves a growth and pro-angiogenic factor.
- An exemplary nucleotide sequence encoding a human MTSP1 is set forth in
SEQ ID Nos - MTSP1 is expressed in breast, prostate and colorectal tumors. Hence conjugates with substrates therefor can be used for treatment of such tumors.
- MTSP3
- The MTSP3 transcript was detected in lung carcinoma (LX-1), colon adenocarcinoma (CX-1), colon adenocarcinoma (GI-112) and ovarian carcinoma (GI-102). No apparent signal was detected in another form of lung carcinoma (GI-117), breast carcinoma (GI-101), pancreatic adenocarcinoma (GI-103) and prostatic adenocarcinoma (PC3).
- MTSP4
- The MTSP4 transcript, a DNA fragment encoding part of the LDL receptor domain and the protease domain was used to probe an RNA blot composed of 76 different human tissues (catalog number 7775-1; human multiple tissue expression (MTE) array; CLONTECH). As in the northern analysis of gel blot, a very strong signal was observed in the liver. Signals in other tissues were observed in (decreasing signal level): fetal liver>heart=kidney=adrenal gland=testis=fetal heart and kidney=skeletal muscle=bladder=placenta>brain=spinal cord=colon=stomach=spleen=lymph node=bone marrow=trachea=uterus=pancreas=salivary gland=mammary gland=lung. MTSP4 also is expressed less abundantly in several tumor cell lines including HeLa S3=leukemia K-562=Burkitt's lymphomas (Raji and Daudi)=colorectal adenocarcinoma (SW480)>lung carcinoma (A549)=leukemia MOLT-4=leukemia HL-60. PCR of the MTSP4 transcript from cDNA libraries made from several human primary tumors xenografted in nude mice (human tumor multiple tissue cDNA panel, catalog number K1522-1, CLONTECH) was performed using MTSP4-specific primers. The MTSP4 transcript was detected in breast carcinoma (GI-101), lung carcinoma (LX-1), colon adenocarcinoma (GI-112) and pancreatic adenocarcinoma (GI-103). No apparent signal was detected in another form of lung carcinoma (GI-117), colon adenocarcinoma (CX-1), ovarian carcinoma (GI-102). and prostatic adenocarcinoma (PC3). The MTSP4 transcript was also detected in LNCaP and PC-3 prostate cancer cell lines as well as in HT-1080 human fibrosarcoma cell line.
- MTSP6
- MTSP6 is expressed at high levels in the colon. It also is expressd in the, stomach, trachea, mammary gland, thyroid gland, salivary gland, pituitary gland and pancreas. It is expressed at lower levels in other tissues (see EXAMPLE 6).
- MTSP6 also is expressed in several tumor cell lines including HeLa S3>colorectal adenocarcinoma (SW480)>leukemia MOLT-4>leukemia K-562. In mouse xenograft models, the MTSP6 transcript was strongly detected in lung carcinoma (LX-1), moderately detected in pancreatic adenocarcinoma (GI-103), weakly detected in ovarian carcinoma (GI-102); and weakly detected in colon adenocarcinoma (GI-112 and CX-1), breast carcinoma (GI-101), lung carcinoma (GI-117) and prostatic adenocarcinoma (PC3). The MTSP6 transcript was also detected in breast cancer cell line MDA-MB-231, prostate cancer cell line PC-3, but not in HT-1080 human fibrosarcoma cell line. MTSP6 also is expressed in mammary gland carcinoma cDNA (Clontech). MTSP6 also is over expressed in ovarian tumor cells.
- MTSP7
- The MTSP7 transcript was detected in lung carcinoma (A549 cell line), leukemia (K-562 cell line) and cervical carcinoma (HeLaS3 cell line). MTSP7 is believed to be expressed in lung, colon, prostate, breast, cervical and other tumors.
- MTSP9
- MTSP9 is, for example, expressed in esophageal tumor tissues, in lung carcinoma, in colorectal carcinoma, lymphoma, a cervical carcinoma (HeLaS3) and leukemia cell lines as well as in certain normal cells and tissues. MTSP9 also can be a marker for breast, prostate, cervical and colon cancer.
- MTSP9 is highly expressed in the esophagus and expressed at a low level in many other tissues. The MTSP9 transcript is found in kidney (adult and fetal), spleen (adult and fetal), placenta, liver (adult and fetal), thymus, peripheral blood leukocyte, lung (adult and fetal), pancreas, lymph node, bone marrow, trachea, uterus, prostate, testes, ovary and the gland organs (mammary, adrenal, thyroid, pituitary and salivary). MTSP9 also is expressed in esophagus tumor tissues, in a lung carcinoma and, at a lower level, in a colorectal carcinoma, lymphoma, a cervical carcinoma (HeLaS3) and leukemia cell lines.
- MTSP10
- MTSP10, for example, is expressed in esophageal tumor tissues, in lung carcinoma, prostate cancers, pancreatic and breast cancers and in cell lines as well as in certain normal cells and tissues (see e.g., EXAMPLES for tissue-specific expression profile). The level of activated MTSP10 can be diagnostic of prostate, uterine, lung esophagus, or colon cancer or leukemia or other cancer. The expression and/or activation of MTSP10 on or in the vicinity of a cell or in a bodily fluid in a subject can be a marker for breast, prostate, lung, colon, esophageal and other cancers.
- MTSP10 transcript was detected in pancreas, lung and kidney. MTSP10 transcript was also detected in small intestine Marathon-Ready cDNA (Clontech). The MTSP10 transcript was detected in breast carcinoma (GI-101), lung carcinoma (LX-1 and GI-117), ovarian carcinoma (GI-102), and pancreatic adenocarcinoma (GI-103). The MTSP10 transcript was weakly detected in prostatic adenocarcinoma (PC3). The MTSP10 transcript was also detected in CWR22R prostate tumor grown in nude mice. No apparent signal was detected in two forms of colon adenocarcinomas (GI-112 and CX-1).
- MTSP12
- MTSP12 transcript was detected in pancreas, lung and kidney. MTSP12 transcript was also detected in small intestine Marathon-Ready cDNA (Clontech). The MTSP12 transcript was detected in breast carcinoma (GI-101), lung carcinoma (LX-1 and GI-117), ovarian carcinoma (GI-102), and pancreatic adenocarcinoma (GI-103). The MTSP12 transcript was weakly detected in prostatic adenocarcinoma (PC3). The MTSP12 transcript was also detected in CWR22R prostate tumor grown on nude mice. No apparent signal was detected in two forms of colon adenocarcinomas (GI-112 and CX-1).
- MTSP20
- MTSP20 is expressed in the lung, colon, cervical tumors and in leukemic cells. It may also be expressed in breast, ovarian, pancreatic, prostate and in other tumors. MTSP20 transcript was detected in liver, lymph node, cerebellum, pancreas, prostate, uterus, testis, glands (adrenal, thyroid and salivary), thymus, kidney and spleen. Lower transcript level was found in lung, placenta, bladder, ovary, digestive system, circulatory system and other parts of the the brain. MTSP20 is also expressed in certain tumor cell lines including lung carcinoma (A519), colorectal carcinoma (SW480), lymphoma (Raji and Daudi), cervical carcinoma (HeLaS3) and leukemia (HL-60, K-562 and MOLT-4) cell lines.
- MTSP22
- MTSP22 is expressed in the uterine tissue, thymus, adipose tissue, and lymph node. It may also be expressed in lung, stomach, uterine, breast, ovarian, prostate and in other tumors. MTSP22 transcript was detected in some uterus tissue samples, but not in their matched tumor samples. In one of 42 uterus samples, MTSP22 is expressed in tumor and its metastatic tissues, but not in the normal tissue counterpart. MTSP22 is also expressed in some stomach tumors and lung tumors, but not in their normal tissue counterparts. MTSP22 is also expressed in the normal tissue of a pancreas matched cDNA pair. MTSP22-encoding cDNA was detected in thymus, adipose tissue, and lymph node
- MTSP25
- MTSP25 is expressed in breast, colon, uterine, ovarian, kidney, prostate, testicular cancer tissue. It may also be expressed in lung, stomach, prostate and in other tumors. MTSP25 transcript was expressed weakly in the lymph node. In the cancer profiling array analysis, MTSP25 is highly expressed in prostate samples (in normal and cancer samples). MTSP25 was highly expressed in a kidney tumor sample, but not in its normal tissue counterpart. MTSP25 was also expressed a breast cancer samples, but not in its normal tissue counterpart. MTSP25 was expressed in normal uterus samples, but not in their tumor counterparts. MTSP25 expression was also ovarian cancer samples. Among these three samples, the expression of MTSP25 was also detected in one of the matched normal tissue counterparts. MTSP25 expression was also detected in tumor samples in colon cDNA pairs.
- PCR analysis revealed that MTSP25 cDNA was strongly detected in testis and mammary gland adenocarcinoma, weakly detected in brain, placenta, lung, spleen, prostate, small intestine, colon, and leukocyte, and very weakly detected in heart, liver and pancreas.
- 2. Endotheliases
- Endotheliases are a class of cell surface proteases that are expressed on cells, particularly endothelial cells, particularly those proliferating endothelial cells, which are involved in a variety of proliferative processes, including undesirable angiogenesis associated with tumor growth and metastasis, and with other hyperproliferative disorders, such as restenosis, scarring, diabetic retinopathies, diseases and disorders of the anterior eye (see, U.S. application Ser. No. 09/717,473, filed Nov. 20, 2000, and International PCT application No. PCT/US00/31803).
- Proliferative Diseases
- Endotheliases are particularly useful targets for delivery of therapeutic agents for treatment of any disorder involving aberrant angiogenesis. Endothelial cells play a key role in angiogenesis, which is is the generation of new blood vessels from parent microvessels. Angiogenesis plays a major role in the metastasis of cancer and in the pathology of a variety of other disorders.
- Controlled and uncontrolled angiogenesis proceed in a similar manner. Endothelial cells and pericytes, surrounded by a basement membrane, form capillary blood vessels. Angiogenesis begins with the erosion of the basement membrane by enzymes released by endothelial cells and leukocytes. The endothelial cells, which line the lumen of blood vessels, then protrude through the basement membrane. Angiogenic stimulants induce the endothelial cells to migrate through the eroded basement membrane. The migrating cells form a “sprout” off the parent blood vessel, where the endothelial cells undergo mitosis and proliferate. The endothelial sprouts merge with each other to form capillary loops, creating the new blood vessel.
- Angiogenesis, Modulators and Associated Diseases
- Angiogenesis is highly regulated by a system of angiogenic stimulators and inhibitors. Known examples of angiogenesis stimulators include certain growth factors, cytokines, proteins, peptides, carbohydrates and lipids (Norrby (1997)APMIS 105:417-437); Polverini (1995) Crit. Rev. Oral. Biol. Med. 6:230-247). A variety of endogenous and exogenous angiogenesis inhibitors are known in the art (Jackson et al. (1997) FASEB 11:457-465; Norrby (1997) APMIS 105:417-437); and O'Reilly (1997) Investigational New Drugs, 15:5-13).
- Angiogenesis is essential for normal placental, embryonic, fetal and post-natal development and growth, but almost never occurs physiologically in adulthood except in very specific restricted situations. For example, angiogenesis is normally observed in wound healing, fetal and embryonal development and formation of the corpus luteum, endometrium and placenta. Angiogenesis in the adult is often associated with disease states.
- Persistent, unregulated angiogenesis occurs in a multiplicity of disease states, tumor metastasis and abnormal growth by endothelial cells and supports the pathological damage seen in these conditions. The diverse pathological disease states in which unregulated angiogenesis is present have been grouped together as angiogenic dependent or angiogenic associated diseases.
- The control of angiogenesis is altered in certain disease states and, in many cases, the pathological damage associated with the disease is related to uncontrolled angiogenesis (see generally, Norrby (1997)APMIS 105:417-437); and O'Reilly (1997) Investigational New Drugs 15:5-13). Thus, angiogenesis is involved in the manifestation or progress of various diseases, for example, various inflammatory diseases, such as rheumatoid arthritis, psoriasis, diabetic retinopathies, certain ocular disorders, including recurrence of pterygii, scarring excimer laser surgery and glaucoma filtering surgery, various disorders of the anterior eye, cardiovascular disorders, chronic inflammatory diseases, wound repair, circulatory disorders, crest syndromes, dermatological disorders (see, e.g., U.S. Pat. Nos. 5,593,990, 5,629,327 and 5,712,291) and notably cancer, including solid neoplasms and vascular tumors. Angiogenesis is essential for the growth and persistence of solid tumors and their metastases. Repressing, eliminating or modulating this activity, should impact the etiology of these diseases and serve as a point of therapeutic intervention. In the disease state, prevention of angiogenesis could avert the damage caused by the invasion of the new microvascular system. Therapies directed at control of the angiogenic processes could lead to the abrogation or mitigation of these diseases. Hence there is a need to develop therapeutics that target angiogenesis and modulate, particularly, inhibit aberrant or uncontrolled angiogenesis.
- Hence conjugates that contain endotheliase substrates can be used to deliver therapeutic agents for the treatment of diseases including, but are not limited to, rheumatoid arthritis, psoriasis, diabetic retinopathies, other ocular disorders, including recurrence of pterygii, scarring from excimer laser surgery and glaucoma filtering surgery, various disorders of the anterior eye, cardiovascular disorders, autoimmune diseases, chronic inflammatory diseases, wounds, circulatory disorders, crest syndromes, restenosis, psoriasis and other dermatological disorders (see, e.g., U.S. Pat. Nos. 5,593,990, 5,629,327 and 5,712,291) and notably cancer, including solid neoplasms and vascular tumors.
- Endotheliases 1 and 2
- Exemplary of endotheliases are two different endotheliases and variant forms thereof designated endotheliase 1 and endotheliase 2 (see SEQ ID Nos. 21-27. Other members of the family can be identified by probing for genes or searching libraries for genes that have sequence identity, particularly at least 40%, 60%, 80%, 90%, 95%, 98% or greater sequence identity to the protease domain of an endotheliase identified herein, or that hybridize under conditions of high stringency to the full-length of the nucleic acid encoding a protease domain of an endotheliase provided herein, and that are expressed on endothelial cells.
- Alternatively, and as a way of identifying endotheliases that can have lower sequence identity, an endotheliase can be identified by the methods, such by identifying ESTs or other nucleic acid fragments that have sequences similar to a protease and then using such fragments as probes to identify and select cDNA clones encoding full-length proteases or protease domains thereof, identifying those that have the characteristics of transmembrane proteins, and then determining the gene expression profile to identify those that are expressed on the surface of endothelial cells. Encoded proteins that have protease activity, that include a transmembrane domain and an extracellular domain, and that are expressed in endothelial cells are endotheliases. Any method for identification of genes encoding proteins (or proteins) that encode a transmembrane protease expressed on an endothelial cell is contemplated herein.
- Endotheliase 1
- Exemplary of the endotheliase are endotheliase 1 and
endotheliase 2. These are expressed on endothelial cells. Exemplary of a full-length endotheliase 1 is one that includes the sequence of amino acids set forth in SEQ ID No. 42 (see, International PCT application No. WO 00/5006, which describes a gene it designates DESC1 that is expressed in squamous cell carcinomas and prostate tumors). As noted endotheliases are expressed on endothelial cells. A protease domain thereof is set forth in SEQ ID NO: 22. - Expression Profile of
Endotheliase 1 - To obtain information regarding the tissue distribution of
endotheliase 1, the DNA insert of clone H117 was used to probe an RNA blot composed of 76 different human tissues (catalog number 7775-1; human multiple tissue expression (MTE) array; CLONTECH, Palo Alto, Calif.). Significant expression was observed in the esophagus, with minor expression levels in the stomach, salivary gland, pancreas, prostate, bladder, trachea and uterus. Northern analysis using RNA blots (catalog numbers 7765-1 & 7782-1; human muscle and digestive system multiple tissue northern (MTN) blots; CLONTECH) confirmed that the expression was restricted to the esophagus. Two transcripts (approximately 1.7 and 2 kb) were detected in the esophagus. Endotheliase 1 also is expressed in umbilical vein endothelial cells, PC3 and LnCAP cells. - Endotheliase 2 and Nucleic
Acids Encoding Endotheliase 2 - Two splice variant forms of
endotheliase 2 designated endotheliase 2-S and endotheliase 2-L are exemplified herein (see SEQ ID Nos. 23-26). The open reading frame of the nucleic acid encoding endotheliase 2-S (SEQ ID No. 23) is composed of 1,689 bp, which translates to a 562-amino acid protein (SEQ ID No. 24), while the ORF of endotheliase 2-L is composed of 2,067 bp (SEQ ID No. 25), which translates to a 688-amino acid protein (SEQ ID No. 26). - The nucleic acid encoding the protease domain of endotheliase 2-S is composed of 729 bp which translates to a 242-amino acid protein (amino acids 321-562 of SEQ ID Nos. 23 and 24), while that of endotheliase 2-L is composed of 1,107 bp, which translates to a 368-amino acid protein (amino acids 321-688 of SEQ ID Nos. 25 and 26).
- Endotheliase-2 Proteins
- Any and all of the above-noted endotheliases and/or protease domains thereof, such as those that include the sequences of amino acids in SEQ ID Nos. 22, 24, 26 and 27 or are encoded by nucleic acid that hybridize thereto under the conditions as described above are contemplated for use in the methods herein. Also contemplated herein are proteins that include amino acid sequence changes, such as those set forth in Table 1 above, and retain protease activity.
- Gene Expression Profile and Transcript Size of
Endotheliase 2 in Normal and Tumor Tissues - In addition to expression in endothelial cells, endotheliase 2 is expressed in placenta, pancreas, thyroid gland, liver and lung tissues. It also is expressed at lower levels in mammary gland, salivary gland, kidney, trachea, esophagus, appendix, heart and fetal lung. Endotheliase 2 also is expressed in several tumor cell lines and, hence, in certain tumors, including lung and colon, including breast carcinoma, lung carcinomas, colon adenocarcinomas, pancreatic adenocarcinoma (GI-103), and ovarian carcinoma. It has also been detected in prostate and fibrosarcoma cell lines.
- Conjugates that are substrates for proteases on the surfaces of cells, particularly serine proteases, including type II membrane-bound serine proteases, and endotheliases are provided. Any cell surface protease, including cell-associated or localized proteases, is contemplated herein. Generally proteases expressed at high levels in active forms in essential tissues are not ideal target candidates. The proteases include those that are expressed on relatively limited numbers of cells or that are expressed at high levels in cells, such as tumor cells and endothelial cells and immune cells, that are involved in disease states or are present in diseases states in the locale of cells involved in the disease states. For example, endothelial cells by virtue of their role in angiogenesis are involved in numerous proliferative disorders; immune cells are involved in many disease processes including cancers and diseases and inflammatory disorders. Other cell surface proteases are expressed at higher levels in certain tumors than in normal cells. Whether or not such proteases have a role in the disorder their higher expression in cells involved in a disease state is sufficient for use for targeting therapeutic agents in the conjugates provided herein.
- The conjugates, which contain a therapeutic agent, such as a cytotoxic agent, is activated upon cleavage by a cell surface protease, including cell-associated and cell-localized proteses. Exemplary of such proteases are the MTSPs, such as, but not limited to, MTSP1, MTSP3, MTsP4, MTSP6, MTSP7, MTSP9, MTSP10, MTSP12, MTSP20, MTSP22, MTSP25, urokinases and endotheliases. Hence, the conjugates targeted to such proteases are prodrugs in that the therapeutic agent is inactive as administered and is ultimately activated in the vicinity of the targeted cell or tissue. Although cell surface proteases, such as transmembrane proteases, are the intended targets, any released, shed or soluble forms of the proteases and others also can be targeted.
- Thus, the conjugates, which contain a therapeutic agent, such as a cytotoxic agent, are substantially inactive prior to action by a cell surface protease, a peptidic moiety that is a substrate for a targeted cell surface protease (i.e., a peptidic substrate), and, optionally, a linker. The therapeutic agents in the conjugates are activated upon cleavage of the peptidic substrate of the conjugate by a cell surface protease. The therapeutic agents, such as cytotoxic agents, are released as the free yagent, or, alternatively, are released coupled to the portion of the peptidic substrate (P1-P2-P3-etc. (i.e., the N-terminus) or P1′-P2′-etc. (i.e., the C-terminus) that the agents were linked to in the conjugate, optionally via a linker. The cytotoxic agents, in these forms, are released in the vicinity of cells that express the proteases. Activation is effected, in certain embodiments, because the therapeutic agent, such as cytotoxic agent, following action of the cell surface protease, can cross the cell membrane or otherwise interact with the cell or tissue and exhibit therapeutic activity. In other embodiments, any remaining peptidic moieties or amino acids can be cleaved from therapeutic agent to render it active. The conjugates act as prodrugs because the therapeutic agents when conjugated are substantially inactive. Upon cleavage by the targeted protease, the therapeutic agent is released either in active form or in a form that is activated by the targeted cell, tissue or surrounding environment.
- In one exemplary embodiment, the targeted agent is a cytotoxic agent and the conjugates for use in the methods and compositions provided herein have the formula:
- (peptidel)s-(linker)q-(cytotoxic agent)t
- or a derivative thereof, where peptidel is a peptidic substrate for a cell surface protease or a released, shed or otherwise unbound membrane protease, such as an MTSP; s is greater than or equal to 1, or is 1 to 6, or is 1 or 2, or is 1; linker is any linker; q is greater than or equal to 0, or is 0 to 4, or is 0 or 1; the cytotoxic agent is an anti-tumor, anti-cancer or anti mitotic agent, including anti-antiangiogenic agents; and t is 1 or more, or is 1 or 2. In these conjugates, the cytotoxic agent is covalently attached, optionally via a linker, to either the C-terminus or the N-terminus of the peptidic substrate. In embodiments where the therapeutic agent, such as a cytotoxic agent, is attached to the C-terminus of the peptidic substrate, the N-terminus optionally is capped. N-Terminal caps for use herein include, but are not limited to, acyl, sulfonyl and carbamoyl groups. In embodiments where the therapeutic agent is attached to the N-terminus of the peptidic substrate, the C-terminus is a carboxamide derivative.
- In certain embodiments, peptidel is a peptidic substrate for a cell surface protease or a soluble MTSP whereby, upon action of the protease, the conjugate, which is substantially inactive, is cleaved at the P1-P1′ bond to release a compound of the formula:
- (peptidea)s-(linker)q-(therapeutic agent)t
- or a derivative thereof, that exhibits therapeutic activity, such as cytotoxic activity in vitro and in vivo. In these compounds, peptidea is a truncated version of peptidel resulting from cleavage at the P1-P1′ bond.
- In another embodiment, the conjugates for use in the methods and compositions provided herein possess two therapeutic agents, such as cytotoxic agents, which are the same or different, linked to the C-terminus and the N-terminus, respectively, optionally via linkers linker1 and linker2, of a peptidic substrate for cell surface protease or a soluble MTSP. In this embodiment, the conjugates have the formula:
- (therapeutic agent1)x-(linker1)w-(peptidel)s-(linker2)q-(therapeutic agent2)t
- or a derivative thereof, where peptidel is a peptidic substrate for a cell surface protease, or a soluble MTSP; s is greater than or equal to 1, or is 1 to 6, or is 1 or 2, or is 1; linker1 and linker2 are each independently any linker and are the same or different; q and w are each independently greater than or equal to 0, or are 0 to 4, or are 0 or 1; the therapeutic agents, which are the same or different, are anti-tumor, anti-cancer or anti mitotic agents; and t and x are each independently 1 or more, or are 1 or 2.
- In these embodiments, peptidel is a peptidic substrate for a cell surface protease or a soluble MTSP whereby, upon action of the protease, the conjugate, which is substantially inactive, is cleaved at a point on the peptidic chain to release two compounds of the formulae:
- (therapeutic agent1)x-(linker1)w-(peptidea1)s; and
- (peptidea2)s-(linker2)q-(therapeutic agent2)t
- or derivatives thereof. The released therapeutic agents are active or are further activated by the cell, tissue or surrounding environment. In these compounds, peptidea1 and peptidea2 are N-terminal and C-terminal truncated portions, respectively, of peptidel resulting from cleavage at the P1-P1′ bond.
- In one embodiment, the conjugates for use in the compositions and methods provided herein have formula I:
- Xn-(P6)m-(P5)p-(P4)i-(P3)j-(P2)l-P1-(P1′)u-(P2′)k-(P3′)r-(L)n-Z
- or a derivative thereof, where Z is a therapeutic agent; L is a linker; l, j, i, p and m are selected as follows:
- l is 0 or 1; when l is 0, j, i, p and m are 0; when l is 1, j is 0 or 1; when j is 0, i, p and m are 0; when j is 1, i is 0 or 1; when i is 0, p and m are 0; when i is 1, p is 0 or 1; when p is 0, m is 0; when p is 1, m is 0 or 1;
- u, k and r are selected as follows:
- u is 0 or 1; when u is 0, k and r are 0; when u is 1, k is 0 or 1; when k is 0, r is 0; when k is 1, r is 0 or 1;
- n is 0 or 1; Xn is hydrogen, or an acyl, sulfonyl or carbamoyl cap; and P6 to P3′ are amino acid residues, as defined below. In this embodiment, the P6 to P3′ residues are linked by peptide bonds or peptide bond surrogates. Thus, the P6 to P3′ portion of the conjugate is a peptidic substrate, as defined herein.
- In another embodiment, the conjugates for use in the compositions and methods provided herein have formula II:
- Z-(L)n-(P6)m-(P5)p-(P4)i-(P3)j-(P2)l-P1-(P1′)u-(P2′)k-(P3′)r-Xc
- or a derivative thereof, where Z is a therapeutic agent; L is a linker; l, j, i, p and m are selected as follows:
- l is 0 or 1; when l is 0, j, i, p and m are 0; when l is 1, j is 0 or 1; when j is 0, i, p and m are 0; when j is 1, i is 0 or 1; when i is 0, p and m are 0; when i is 1, p is 0 or 1; when p is 0, m is 0; when p is 1, m is 0 or 1;
- u, k and r are selected as follows:
- u is 0 or 1; when u is 0, k and r are 0; when u is 1, k is 0 or 1; when k is 0, r is 0; when k is 1, r is 0 or 1;
- n is 0 or 1; Xc, together with the carbonyl group of the amino acid residue to which it is attached, forms a carboxylic acid or a carboxamide group; and P6 to P3′ are amino acid residues, as defined below. In this embodiment, the P6 to P3′ residues are linked by peptide bonds or peptide bond surrogates. Thus, the P6 to P3′ portion of the conjugate is a peptidic substrate, as defined herein.
- In a further embodiment, the conjugates for use in the compositions and methods provided herein have formula III:
- Z1-(L1)n-(P6)m-(P5)p-(P4)i-(P3)j-(P2)l-P1-(P1′)u-(P2′)k-(P3′)r(L2)v-Z2
- or a derivative thereof, where Z1 and Z2 are each therapeutic agents and are the same or different; L1 and L2 are each linkers and are the same or different; l, j, i, p and m are selected as follows:
- l is 0 or 1; when l is 0, j, i, p and m are 0; when l is 1, j is 0 or 1; when j is 0, i, p and m are 0; when j is 1, i is 0 or 1; when i is 0, p and m are 0; when i is 1, p is 0 or 1; when p is 0, m is 0; when p is 1, m is 0 or 1;
- u, k and r are selected as follows:
- u is 0 or 1; when u is 0, k and r are 0; when u is 1, k is 0 or 1; when k is 0, r is 0; when k is 1, r is 0 or 1;
- n and v are each independently 0 or 1; and P6 to P3′ are amino acid residues, as defined below. In this embodiment, the P6 to P3′ residues are linked by peptide bonds or peptide bond surrogates. Thus, the P6 to P3′ portion of the conjugate is a peptidic substrate, as defined herein.
- In another embodiment, the conjugates for use in the compositions and methods provided herein have formula IV:
- Xn-(P6)m-(P5)p-(P4)i-(P3)j-(P2)l-P1-(P1′)u-(P2′)k-(P3′)r-(P4′)s-(L)n-Z
- or a derivative thereof, where Z is a therapeutic agent; L is a linker; l, j, i, p and m are selected as follows:
- l is 0 or 1; when l is 0, j, i, p and m are 0; when l is 1, j is 0 or 1; when j is 0, i, p and m are 0; when j is 1, i is 0 or 1; when i is 0, p and m are 0; when i is 1, p is 0 or 1; when p is 0, m is 0; when p is 1, m is 0 or 1;
- u, k, r and s are selected as follows:
- u is 0 or 1; when u is 0, k, r and s are 0; when u is 1, k is 0 or 1; when k is 0, r and s are 0; when k is 1, r is 0 or 1; when r is 0, s is 0; when r is 1, s is 0 or 1;
- n is 0 or 1; Xn is hydrogen, or an acyl, sulfonyl or carbamoyl cap; and P6 to P4′ are amino acid residues, as defined below. In this embodiment, the P6 to P4′ residues are linked by peptide bonds or peptide bond surrogates. Thus, the P6 to P4′ portion of the conjugate is a peptidic substrate, as defined herein. In another embodiment, the conjugates for use in the compositions and methods provided herein have formula V:
- Z-(L)n-(P6)m-(P5)p-(P4)i-(P3)j-(P2)l-P1-(P1′)u-(P2′)k-(P3′)r-(P4′)s-Xc
- or a derivative thereof, where Z is a therapeutic agent; L is a linker; i, j, i, p and m are selected as follows:
- l is 0 or 1; when l is 0, j, i, p and m are 0; when l is 1, j is 0 or 1; when j is 0, i, p and m are 0; when j is 1, i is 0 or 1; when i is 0, p and m are 0; when i is 1, p is 0 or 1; when p is 0, m is 0; when p is 1, m is 0 or 1;
- u, k, r and s are selected as follows:
- u is 0 or 1; when u is 0, k, r and s are 0; when u is 1, k is 0 or 1; when k is 0, r and s are 0; when k is 1, r is 0 or 1; when r is 0, s is 0; when r is 1, s is 0 or 1;
- n is 0 or 1; Xc, together with the carbonyl group of the amino acid residue to which it is attached, forms a carboxylic acid or a carboxamide group; and P6 to P4′ are amino acid residues, as defined below. In this embodiment, the P6 to P4′ residues are linked by peptide bonds or peptide bond surrogates. Thus, the P6 to P4′ portion of the conjugate is a peptidic substrate, as defined herein.
- In a further embodiment, the conjugates for use in the compositions and methods provided herein have formula VI:
- Z1-(L1)n-(P6)m-(P5)p-(P4)i-(P3)j-(P2)l-P1-(P1′)u-(P2′)k-(P3′)r-(P4′)s-(L2)v-Z2
- or a derivative thereof, where Z1 and Z2 are each therapeutic agents and are the same or different; L1 and L2 are each linkers and are the same or different; l, j, i, p and m are selected as follows:
- l is 0 or 1; when l is 0, j, i, p and m are 0; when l is 1, j is 0 or 1; when j is 0, i, p and m are 0; when j is 1, i is 0 or 1; when i is 0, p and m are 0; when i is 1, p is 0 or 1; when p is 0, m is 0; when p is 1, m is 0 or 1;
- u, k, r and s are selected as follows:
- u is 0 or 1; when u is 0, k, r and s are 0; when u is 1, k is 0 or 1; when k is 0, r and s are 0; when k is 1, r is 0 or 1; when r is 0, s is 0; when r is 1, s is 0 or 1;
- n and v are each independently 0 or 1; and P6 to P4′ are amino acid residues, as defined below. In this embodiment, the P6 to P4′ residues are linked by peptide bonds or peptide bond surrogates. Thus, the P6 to P4′ portion of the conjugate is a peptidic substrate, as defined herein.
- Exemplary peptidic substrates, therapeutic agents, linkers and exemplary conjugates of formulae I-VI are described in further detail below. It is intended herein that conjugates resulting from all combinations and/or permutations of the groups recited below for the variables of formulae I-VI are encompassed within the instant disclosure.
-
- The peptidic substrates contemplated for use in the conjugates are substrates for the targeted cell surface protease or a soluble, shed or released form thereof, and contain a sufficient number of amino acid residues to render any therapeutic agent in the conjugate substantially inactive. In the exemplary embodiment where the therapeutic agent is, for example, doxorubicin, the conjugate is substantially inactive by virtue of the inability of the conjugated therapeutic agent to cross the cell membrane. In certain embodiments, the peptidic substrate contains at least 1, 2, 3, 4 or 5 amino acid residues, and can contain up to nine or ten residues. Longer peptidic substrates can be used in the conjugates as long as upon cleavage, the resulting therapeutic agent or therapeutic agent-amino acid or -peptidic moiety conjugate exhibits the desired therapeutic effect in vivo and in vitro.
- Hence, exemplary peptidic substrates for use in the conjugates provided herein possess at least one amino acid (P1), two amino acids (P1-P1′), three amino acids (P2-P1-P1′) and typically contain four, five or six amino acid residues (P3-P2-P1-P1′, P4-P3-P2-P1-P1′ or P4-P3-P2-P1-P1′-P2′), where the P1-P1′ bond is the site of cleavage of cell surface protease, or a soluble, shed or released form thereof, including, but not limited to, a cell surface protease, such as a serine protease, including, for example, but not limited to, uPA bound to its receptor, MTSPs and endotheliases. The peptidic substrates optionally further possess a P5, P6 or P3′ amino acid residue, and, in certain embodiments, possess P7, P8, P9, P10, P4′, P5′, P6′ residues. Thus, the peptidic substrates for use in the conjugates provided herein are penta-, hexa-, hepta-, octa- and nona-peptidic substrates, and can contain 10, 11, 12, 13, 14, 15 or more residues as long as, upon cleavage of the conjugate by the protease, the resulting therapeutic agent or therapeutic agent-amino acid or -peptidic moiety conjugate exhibits the desired therapeutic effect in vivo and in vitro.
- The peptidic substrates are conjugated to the therapeutic agent (or to a linker to which the therapeutic agent is linked) via the C-terminal residue (i.e., P1′, P2′ or P3′), or the N-terminal residue (i.e., P6, P5 or P4), or optionally an internal residue. The peptidic substrates, for example, can be straight chains, but can be cyclized or include cyclized portions.
- In embodiments where the conjugation is via the C-terminus of the peptidic substrate, the peptidic substrate optionally possesses a cap, such as an acyl or carbamoyl cap at the N-terminus. In embodiments where conjugation is via the N-terminus of the peptidic substrate, the peptidic substrate further possess a terminal group, such as a carboxamide group, at the C-terminus.
- The conjugates can contain a plurality of peptidic substrates and a plurality of therapeutic agents. For example, in conjugates that contain two therapeutic agents, which are the same or different, conjugation to the therapeutic agent(s) or linker linked thereto can be via the C-terminal and N-terminal residues of the peptidic substrate.
- The methods described for selection of substrates above can be used to design suitable substrates. In addition, substrates can be designed based upon known specificities of other proteases. For example, the specificities of trypsin-like and trypsin family members can aid in design of possible substrates. The following summarizes substrate preferences for particular serine proteases (see, e.g., Harris et al. (2000)PNAS 97(14):7754-7759).
EXEMPLARY EXEMPLARY EXEMPLARY PROTEASE P1 RESIDUE(S) P2 RESIDUE(S) P3 RESIDUE(S) Chymotrypsin Tyr, Phe, Trp — — Trypsin Arg, Lys — — Thrombin Arg, Lys Phe Thr, Trp Plasmin Lys, Arg Trp, Tyr, Met Gln Granzyme B Asp — — Human Ala, Val, Ile — — Neutrophil Elastase Tissue Arg Ser, Gly, Ala Met, Tyr Plasminogen Factor Urokinase Arg Ser, Ala Thr, Ser Factor Xa Arg Gly — - Typical protocols for preparation of the conjugates can include the steps of: 1) identification of a targeted protease; 2) expression and assay development; 3) substrate selection, such as, for example, by testing chromogenic or fluorogenic substrates to identify those cleaved by a selected target protease, by use of substrate phage display to identify peptidic substrates cleaved by a targeted protease, by use of a natural protein or peptide substrate or a natural inhibitor of the protease, and by use of combinatorial libraries to identify substrates cleaved by a targeted protease; 4) synthesis of conjugates containing the identified substrate; and 5) biological evaluation thereof, including, but not limited to, in vitro assays, cell culture assays, biological assays, and in vivo animal models (see, e.g., EXAMPLE 10).
- A conjugate can be designed by any methods known to those of skill in the art. The following provides an exemplary protocol. First, a series of commercially available chromogenic and fluorogenic peptidic substrates can be tested for cleavage by the protease of interest (see Examples for lists of exemplary chromogenic and fluorogenic substrates and the table below). The peptidic portion of these substrates occupies the unprimed binding sites of the protease while the reporter group is located on the primed side of the scissle bond. Effective conjugates can then be designed based on the structure of the substrates that are efficiently cleaved by the protease.
- The peptidic portion of these efficiently cleaved substrates can be used as the unprimed region of the conjugate, and Ser-therapeutic agent, such as a cytotoxic agent (e.g., doxorubicin), Ser-Leu-therapeutic agent or Ser-Ser-Leu-therapeutic agent can be used as the primed region of the conjugate. Cleavage of these conjugate prodrugs releases either Ser-therapeutic agent, Ser-Leu-therapeutic agent or Ser-Ser-Leu-therapeutic agent compounds. In another embodiment, the Ser in the released Ser-therapeutic agent may be replaced by other amino acid residues including, but not limited to, Ala, hSer, Abu, Thr, Met, nLeu and Val. In another embodiment, such as when the therapeutic agent is doxorubicin, the amino acid residue conjugated to the therapeutic agent possesses a hydrophobic side chain. Such amino acid residues include, but are not limited to, Leu, Abu, nLeu, nVal, CHA, hCHA, (hex)Gly, (allyl)Gly, (propargyl)Gly and (cyclopropyl)Ala. In another embodiment, such as when the therapeutic agent is taxol, the amino acid residue conjugated to the therapeutic agent possesses a side chain that is not sterically bulky. Such amino acid residues include, but are not limited to, Gly and Ala. The resulting P1′-therapeutic agent, P1′-P2′-therapeutic agent, or P1′-P2′-P3′-therapeutic agent compound can be further processed in vivo into active therapeutic agents.
- Another approach to designing a conjugate prodrug for a protease substrate is to use substrate phage display to elucidate optimal subsite occupancy for the protease. The resulting information can then be used to design the peptidic, unprimed portion of the conjugate. As described above, the primed region of the conjugate can be fixed as Ser-therapeutic agent, Ser-Leu-therapeutic agent or Ser-Ser-Leu-therapeutic agent.
- A third approach to design an effective prodrug conjugate involves the use of combinatorial fluorogenic substrate libraries to determine optimal residues for the unprimed region of a protease substrate. These selected sequences can then be used as the unprimed portion of the conjugate prodrug and, and Ser-therapeutic agent, (e.g., doxorubicin), Ser-Leu-therapeutic agent or Ser-Ser-Leu-therapeutic agent can be used as the primed region of the conjugate. These methods have been used in the design of the peptidic substrate portion of the conjugates provided herein. For example, sequences including GSGR (and related sequences such as TGR, SGR, extended variants and others herein) were based on or dervied from substrate phage display experiments using u-PA as the taret protease. Many matriptase conjugates, such as (R/K)-X-S-R and X-(R/K)-S-R, and related sequences as provided herein, were based on data from combinatorial libraries. In other embodiemnts, seqeuence sequences in natural substrates or natural inhibitors of a protease target, such as uPA, including VSAR, PGR (from P3-P1 of plasminogen) and related sequences, were used in design of u-PA-targetd conjugates. In other embodiments, sequences from chromgenic substrates, such as D-HHT-Gly-Arg, and related sequences, were used for design of ET-1-targeted conjugates.
- Chromogenic/Fluorogenic Substrates
Chromogenic/fluorogenic substrates Enzyme Substrate Structure MTSP1 Spectrozyme t-PA CH3SO2-D-HHT-Gly-Arg-pNA.AcOH MTSP1 S 2765 N-α-Z-D-Arg-Gly-Arg-pNA.2HCl MTSP3 Spectrozyme fXIIa H-D-CHT-Gly-Arg-pNA.2AcOH MTSP4a Spec PL H-D-Nle-HHT-Lys-pNA.2AcOH MTSP5 S 2765 N-α-Z-D-Arg-Gly-Arg-pNA.2HCl MTSP6 spectrozyme t-PA CH3SO2-D-HHT-Gly-Arg-pNA.AcOH MTSP7 S 2366 pyroGlu-Pro-Arg-pNA.HCl MTSP9 Pefachrome fVIIa CH3SO2-D-CHA-But-Arg-pNA MTSP10 spectrozyme t-PA CH3SO2-D-HHT-Gly-Arg-pNA.AcOH MTSP22 S 2366 pyroGlu-Pro-Arg-pNA.HCl ET-1 spectrozyme t-PA CH3SO2-D-HHT-Gly-Arg-pNA.AcOH ET-2 S 2765 N-α-Z-D-Arg-Gly-Arg-pNA.2HCl u-PA S-2444 pyroGlu-Gly-Arg-pNA.HCl - Briefly, for a coupled assay, the ability of the protease to activate an enzyme, such as plasminogen or trypsinogen is tested. To perform these assays, a protease is incubated with a zymogen, such as plasminogen or trypsinogen, in the presence of a labelled known substrate, such as lys-plasminogen or Spec PL (for plasmin), for the zymogen. If protease activates the zymogen, the activated enzyme, such as plasmin and trypsin, will degrade the substrate, thereby changing the spectral properties of the substrate.
- Exemplary Peptidic Substrates
- The following description provides exemplary peptidic substrates for cleavage by proteases, such as MTSP1 (or matriptase),
endotheliase 1 and urokinase, and a general discussion of properties of the residues. In a similar manner, peptidic substrates for cleavage by other cell surface proteases, or a soluble, shed or released form thereof, can be similarly designed by identifying peptidic substrates for the selected protease and then preparing conjugates that contain such peptidic substrates. - a. The P1 Residue
- Amino acid residues for use at the P1 position of the peptidic substrates for use in the conjugates provided herein include Arg, Arg surrogates and Lys. Arg surrogates include unnatural amino acids that possess a group or moiety that functions in substantially the same way as the naturally occurring side chain of arginine to achieve substantially the same result (e.g., acting as the P1 residue in a substrate for a MTSP1, urokinase or endotheliase). Arg surrogates include, but are not limited to, α-amino acids that possess as the side chain any of the following: the side chain of homoarginine; guanidinoaminopropyl; guanidinoaminoethyl; (Me)2arginine side chain; (Et)2arginine side chain; (4-aminomethyl)phenylmethyl; 4-amidinophenylmethyl; 4-guanidinophenylmethyl; or the Arg surrogate is a conformationally constrained arginine analog such as:
-
- where d is an integer from 0 to 5, or 1 to 3; and W is N or CH; or a mono- or di-substituted N-alkyl derivative of the above groups, where alkyl is, in certain embodiments, lower alkyl, such as, for example, methyl.
- In certain embodiments herein, the P1 residue is Arg.
- b. The P2 Residue
- In the conjugates provided herein, the P2 residue is selected from Phe, Ser, Gly, Ala, Ser(OMe), hSer, 1-methylHis, 3-methylHis, His, nVal, nLeu, Abu, (hS)Gly, Thr, Aib, CHA and Tyr. In another embodiment, the P2 residue is selected from Phe, Ser, Gly and Ala. In certain embodiments herein, the P2 residue is Ser or Ala. In another embodiment, the P2 residue is Gly or Ala.
- c. The P3 Residue
- Amino acid residues for use at the P3 position of the conjugates provided herein include Arg, Lys, Gln, Quat, Arg surrogates, Ser, Thr, hSer, dSer, Pro, (hS)Gly, Tyr, 4,4-dimethylThr, Asn, Met(O2), Quat2, Quat3, Quat4 and Quat5. In another embodiment, the P3 residue is selected from Arg, Lys, Gln, Quat and Arg surrogates. Arg surrogates include those described above for the P1 residue.
- In certain embodiments, the P3 residue is Gin or Ser.
- d. The P4 Residue
- In the conjugates provided for use in the compositions and methods provided herein, the P4 residue is selected from Pro, Arg, Ser, Ala, Lys, Gly, nLeu, Leu, Tyr, Glu, Phe, Val, N,N-dimethylGly, β-Ala, Cys(Me), Gin, t-butylGly and nVal. In another embodiment, the P4 residue is selected from Pro, Arg, Ser, Ala, Lys, Gly, nLeu, Leu, Tyr, Glu, Phe and Val. In further embodiments, the P4 residue is selected from Pro, Arg, Ser, Ala, Lys, Gly, nLeu, Phe or Val. In certain embodiments herein, the P4 residue is Arg or Gly.
- e. The P5 and P6 Residues
- In certain embodiments herein, the peptidic substrates used in the conjugates contain a P5 and, optionally, a P6 residue. P5 residues include Ile, Arg and Arg surrogates. In another embodiment, P5 residues include Arg and Arg surrogates. Arg surrogates include those described above for the P1 residue. P6 residues include, for example, Leu, Val and Arg. In another embodiment, P6 residues include, for example, Leu.
- f. The P1′ Residue
- The P1′ residue of the conjugates provided herein is Gly, Ser, Ala, Leu, Ile, d-Ile, nLeu, Val, nVal, Aib, Abu, Met, 6-aminohexanoyl, Thr or hSer. In another embodiment, the P1′ residue of the conjugates provided herein is Gly, Ser, Ala, Leu, Ile, d-Ile, nLeu, Val, nVal, Aib, Abu, Met or 6-aminohexanoyl. In another embodiment, the P1′ residue is Ser, Ala, hSer, Abu, Thr, Met, nLeu or Val. In another embodiment, the P1′ residue is Gly or Ala. In another embodiment, the P1′ residue is Ser, Ala or Gly. In another embodiment, the P1′ residue is Leu, Abu, nLeu, nVal, CHA, hCHA, (hex)Gly, (allyl)Gly, (propargyl)Gly or (cyclopropyl)Ala. In certain embodiments herein, the P1′ residue is Ala, Ser, Gly, Ile or d-Ile.
- g. The P2′ Residue
- In certain embodiments herein, the conjugates provided herein possess a P2′ residue. P2′ residues for use herein include, but are not limited to, Gly, Ser, Ala, Leu, Ile, d-Ile, nLeu, Val, nVal, Aib, Abu, Met, 6-aminohexanoyl, hCHA, CHA, hexylGly, allylGly and Phe. In another embodiment, P2′ residues for use herein include, but are not limited to, Gly, Ser, Ala, Leu, Ile, d-Ile, nLeu, Val, nVal, Aib, Abu, Met and 6-aminohexanoyl. In another embodiment, the P2′ residue is Ser, hSer, Abu, nLeu, nVal, CHA, hCHA, (allyl)Gly or (hexyl)Gly. In another embodiment, the P2′ residue is Gly or Ala. In another embodiment, the P2′ residue is Leu, Abu, nLeu, nVal, CHA, hCHA, (hex)Gly, (allyl)Gly, (propargyl)Gly or (cyclopropyl)Ala. In further embodiments, the P2′ residues are Ala, Gly, Ile or d-Ile.
- h. The P3′ Residue
- In other embodiments herein, the peptidic substrates used in the conjugates provided herein include a P3′ residue. P3′ residues for use herein include, but are not limited to, Gly, Ser, Ala, Leu, Ile, nLeu, Val, nVal, Aib, Abu, Met, 6-aminohexanoyl, CHA and allylGly. In another embodiment, the P2′ residue is Ser, hSer, Abu, nLeu, nVal, CHA, hCHA, (allyl)Gly or (hexyl)Gly. In another embodiment, P3′ residues for use herein include, but are not limited to, Gly, Ser, Ala, Leu, Ile, nLeu, Val, nVal, Aib, Abu, Met and 6-aminohexanoyl. In another embodiment, the P3′ residue is Gly or Ala. In another embodiment, the P3′ residue is Leu, Abu, nLeu, nVal, CHA, hCHA, (hex)Gly, (allyl)Gly, (propargyl)Gly or (cyclopropyl)Ala.
- i. The P4′ Residue
- In other embodiments herein, the peptidic substrates used in the conjugates provided herein include a P4′ residue. P4′ residues for use herein include, but are not limited to, Gly, Ser, Ala, Leu, Ile, nLeu, Val, nVal, Aib, Abu, Met, 6-aminohexanoyl, CHA and allylGly. In another embodiment, P4′ residues for use herein include, but are not limited to, Gly, Ser, Ala, Leu, Ile, nLeu, Val, nVal, Aib, Abu, Met and 6-aminohexanoyl. In another embodiment, the P4′ residue is Gly or Ala. In another embodiment, the P4′ residue is Leu, Abu, nLeu, nVal, CHA, hCHA, (hex)Gly, (allyl)Gly, (propargyl)Gly or (cyclopropyl)Ala. In another embodiment, the P4′ residue is Leu.
- j. Caps
- 1) Xn (the N-terminal Cap)
- In embodiments herein where the therapeutic agent is conjugated to the C-terminus of the peptidic substrate (i.e., where the conjugate has formula I), the N-terminus of the peptidic substrate optionally is capped with an acyl, sulfonyl or carbamoyl derivative. The cap is chosen, in certain embodiments, to increase the hydrophilicity of the conjugate. In embodiments where the peptidic substrate-therapeutic agent conjugate is sufficiently hydrophilic so as not to require further hydrophilicity, a non-hydrophilic N-terminal cap, such as an acetyl group, can be used. In embodiments where increased hydrophilicity is desired, the N-terminal amino acid is modified with a hydrophilic blocking group. Such blocking groups are chosen based upon the presence of hydrophilic functionality. Such blocking of the terminal amino group can also reduce or eliminate the enzymatic degradation of such peptidyl therapeutic agents by the action of exogenous amino peptidases which are present in the blood plasma of warm blooded animals.
- N-Terminal blocking groups that increase the hydrophilicity of the conjugates and therefore increase the aqueous solubility of the conjugates include, but are not limited to, hydroxylated alkanoyl, polyhydroxylated alkanoyl, polyethylene glycol, glycosylates, sugars and crown ethers.
- In certain embodiments herein, the N-terminal blocking group is one of the following:
-
- where R1 and R2 are selected from (i) or (ii) as follows:
- (i) R1 and R2 are each independently:
- a) hydrogen;
- b) unsubstituted or substituted aryl, unsubstituted or substituted heterocyclyl, C3-C10 cycloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, C1-C6 perfluoroalkyl, R4O—, R3C(O)NR3—, (R3)2NC(O)—, (R3)2N—C(NR3)—, R4S(O)eNH—, —CN, —NO2, R3C(O)—, —N3, —N(R3)2, or R4OC(O)NR3—;
- c) unsubstituted C1-C6 alkyl;
- d) substituted C1-C6 alkyl wherein the substituent on the substituted C1-C6 alkyl is selected from unsubstituted or substituted aryl, unsubstituted or substituted heterocyclyl, C3-C10 cycloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, R3O—, R4S(O)eNH—, R3C(O)NR3—, (R3)2NC(O)—, (R3)2N—C(NR3)—, —CN, R3C(O)—, —N3, —N(R3)2, and R4OC(O)—NR3—; or
- (ii) R1 and R2 are combined to form —(CH2)f— where one of the carbon atoms optionally is replaced by a moiety selected from: —O—, —S(O)e—, —NC(O)—, —NH— and —N(COR4)—;
- R3 is selected from: hydrogen, unsubstituted or substituted aryl, unsubstituted or substituted heterocyclyl, C1-C6 alkyl and C3-C10 cycloalkyl;
- R4 is selected from: unsubstituted or substituted aryl, unsubstituted or substituted heterocyclyl, C1-C6 alkyl and C3-C10 cycloalkyl;
- e is 0, 1 or 2;
- a is 1, 2, 3 or 4;
- b is zero or an integer between 1 and 100; and
- c is 0 to 10, provided that if b is zero, c is 1 to 10; and
- f is 3, 4 or 5.
- In certain embodiments, R1 and R2 are each independently hydrogen, OH, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 aralkyl or aryl. In these embodiments, a is 1, 2, 3 or 4; b is 0 or an integer between 1 and 100; and c is 0 to 10, provided that if b is 0, c is 1 to 10.
-
- where R1 and R2 are each independently hydrogen, C1-C6 alkyl and aryl; a is 1, 2, 3 or 4; a′ is 0, 1, 2 or 3; b is 0 or an integer between 1 and 14; and c is 0 or 1, provided that if b is 0, c is 1.
- In another embodimbent, Xn is R30O—C(O)—, R31R32N—C(O)—, R33(CH2)kC(O)— or H—C(O)—; where k is an integer from 1 to 4, or is 1 or 2; R30 is alkyl, aryl, heteroaryl, aralkyl or heteroaralkyl; R31 and R32 are each independently hydrogen, alkyl, aryl, heteroaryl, aralkyl, or heteroaralkyl; and R33 is hydrogen, hydroxy, alkyl, alkenyl, alkynyl, alkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, aralkyl, aralkoxy, heteroaralkyl or heteroaralkoxy.
- In certain embodiments herein, Xn is hydrogen, acetyl, hydroxyacetyl, 2,3-dihydroxypropionyl, 2,3,4-trihydroxybutanoyl, PEG(1), PEG(2), PEG(4), PEG(6), PEG(14), PEG(15), PEG(16), PEG(17), PEG(18) or PEG(19). In other embodiments herein, Xn is hydrogen, acetyl, hydroxyacetyl, succinyl, quinyl, gallyl, 4-imidazolylacetyl, cotininyl, 3-phosphonylpropionyl, gulonyl, 4-phosphonylbutyryl, glutaryl, ethoxysquaryl or PEG(2). In further embodiments, Xn is hydrogen, acetyl, —C(O)NH2, HOCH2CH2C(O)—, diaminopropanoyl, or NH2—(CH2)5—C(O)—. In another embodiment, Xn is hydrogen, acetyl, succinyl, glutaryl, PEG(2) or malonyl. In another embodiment, Xn is hydrogen, acetyl, succinyl, glutaryl, PEG(2), malonyl, methoxycarbonyl, phenylsulfonyl, 3-methoxypropanoyl, ethoxycarbonyl, isobutoxycarbonyl, benzyloxycarbonyl, tert-butoxycarbonyl, 4-oxopentanoyl, 2-(2-methoxyethoxy)ethoxy)acetyl, 3,4-methylenedioxyphenylacetyl, 2-pyridylacetyl, phenoxyacetyl, phenylacetyl, methoxyacetyl, 2-methoxyethoxycarbonyl, 2-methoxyethoxyacetyl, 3-phenyl-2-hydroxypropanoyl, pent-4-ynoyl, 1-naphthylacetyl, hydroxyacetyl, 3-methoxycarbonylpropanoyl or formyl.
- In certain embodiments herein, the N-terminal cap (Xn) is acetyl, glutaryl, or related acyl, sulfonyl or carbamoyl derivatives. Capping groups include, but are not limited to, a simple N-acetyl residue through larger fragments that impact the overall physicochemical properties of the conjugate. Appropriate choice of the capping group allows delivery of either relatively hydrophilic or hydrophobic molecules to a target site. In one embodiment, Xn is acetyl.
- 2) Xc (the C-terminal Cap)
- In embodiments herein where the therapeutic agent is conjugated to the N-terminus of the peptidic substrate (i.e., where the conjugate has formula II), the C-terminus of the peptidic substrate is a carboxylic acid or a carboxamide derivative. Appropriate choice of the capping group allows delivery of either relatively hydrophilic or hydrophobic molecules to a target site.
- In one embodiment, Xc, together with the carbonyl group to which it is attached, forms a carboxamide derivative of formula —C(O)NRdRe, where Rd and Re are selected from (i) or (ii) as follows:
-
- provided that at least one of Rd and Re are not hydrogen or C1-C6-alkyl; or
- (ii) Rd and Re together form a —CH2CH2OCH2CH2— diradical;
- b is zero or an integer between 1 and 100; and
- c is 0 or 1, provided that if b is zero, c is 1.
- In one embodiment, Rd is hydrogen and Re is 2-hydroxyethyl.
- 2. Linkers
- The conjugates optionally contain a linker (i.e., L, L1 or L2 of formulae I, II and III) that covalently binds the peptidic substrate to the therapeutic agent. The linkers are any that result in a conjugate in which the peptidic portion is a substrate for a cell surface protease and the therapeutic agent is substantially inactive when in the conjugate and is released in active form or in a form subsequently activated by the cell, tissue or environment of the targeted tissue.
- For example, the linker can include of carbohydrate, peptide, diamine, arylamine, and/or hydrocarbon core structures. Linkers are desirably synthetically accessible, provide shelf-stable products, and do not possess any intrinsic biological activity that interferes with the conjugates activity. They can add desirable properties such as increasing solubility or serving to aid in trafficking the cleaved therapeutic agent in the cell. In certain embodiments, some linkers will be enzymatically cleaved in vitro and in vivo, and fragment to release active therapeutic agent or activatable therapeutic agent. In embodiments where the therapeutic agent is doxorubicin, the linker is, for example, a sugar and/or a peptide, such the aminosugar daunosamine.
- In one embodiment, linkers for use herein include, but are not limited to, a biscarbonyl alkyl diradical whereby an amine moiety on the therapeutic agent is connected with the linker unit to form an amide bond and the amino terminus of the peptidic substrate is connected with the other end of the linker unit also forming an amide bond. Conversely, a diaminoalkyl diradical linker unit, whereby a carbonyl moiety on the cytotoxic agent is covalently attached to one of the amines of the linker unit while the other amine of the linker unit is covalently attached to the C-terminus of the peptidic substrate, also can be useful. Other such linker units which are stable to the physiological environment when not in the presence of a cell surface protease, but are cleavable upon the cleavage of the cell surface protease proteolytic cleavage site, are intended for use herein. Furthermore, linker units can be utilized that, upon cleavage of the cell surface protease proteolytic cleavage site, remain attached to the therapeutic agent but do not significantly decrease the therapeutic activity of such a post-cleavage therapeutic agent derivative when compared with an unmodified therapeutic agent.
- In other embodiments, the linker is a diamine containing a cyclic alkyl moiety and, in certain embodiments, the diamine contains a bicycloalkylene moiety. Examples of such diamine linkers include, but are not limited to, 1,4-bis(aminomethyl)cyclohexane, 1,4-bis(aminomethyl)-cycloheptane, 1,3-bis(aminomethyl)cyclopentane, 1-amino-4-(aminomethyl)cyclohexane, 1,4-diaminocyclohexane and 1,4-bis(aminomethyl)-bicyclo[2.2.2]octane.
- Other linkers include 1,ω-diaminoalkanes, including, but not limited to, 1,3-diaminopropane, and 1,ω-dicarbonylalkanes, including, but not limited to, oxalic, malonic, succinic, glutaric, adipic and pivalic acids.
-
- where A is NH or 0; D is N(H or alkyl) or 0; R25 is H, alkyl, cycloalkyl, cycloalkylalkyl, aryl, heteroaryl optionally substituted with 1 or more, such as 1 to 3, substituents selected from halo, halo alkyl and alkyl, aralkyl, heteroaralkyl, alkenyl containing 1 to 2 double bonds, alkynyl containing 1 to 2 triple bonds, alk(en)(yn)yl groups, halo, pseudohalo, cyano, hydroxy, haloalkyl and polyhaloalkyl, such as, for example, halo lower alkyl, especially trifluoromethyl, formyl, alkylcarbonyl, arylcarbonyl that optionally is substituted with 1 or more, such as, for example, 1 to 3, substituents selected from, for example, halo, halo alkyl and alkyl, heteroarylcarbonyl, carboxy, alkoxycarbonyl, aryloxycarbonyl, aminoimino, alkoxycarbonylamino, aryloxycarbonylamino, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, arylaminocarbonyl, diarylaminocarbonyl, aralkylaminocarbonyl, alkoxy, aryloxy, perfluoroalkoxy, alkenyloxy, alkynyloxy, arylalkoxy, aminoalkyl, alkylaminoalkyl, dialkylaminoalkyl, arylaminoalkyl, amino, alkylamino, dialkylamino, arylamino, alkylarylamino, alkylcarbonylamino, arylcarbonylamino, azido, nitro, mercapto, alkylthio, arylthio, perfluoroalkylthio, thiocyano, isothiocyano, alkylsulfinyl, alkylsulfonyl, arylsulfinyl, arylsulfonyl, aminosulfonyl, alkylaminosulfonyl, dialkylaminosulfonyl and arylamino-sulfonyl.; and y is an integer from 1 to 3.
- 3. Therapeutic Agents
- The conjugates are intended for modifying a variety of biological responses. Accordingly, the therapeutic agents are any agents, including proteins and polypeptides, small molecules and other molecules that possess or potentiate a desired biological activity. Such molecules include cytotoxic agents, such as, but are not limited to, a toxin such as abrin, ricin A, pseudomonas exotoxin, shiga toxin, diphtheria toxin and other such toxins and toxic portions and/or subunits or chains thereof; proteins such as, but not limited to, tumor necrosis factor, α-interferon, γ-interferon, nerve growth factor, platelet derived growth factor, tissue plasminogen activator; or, biological response modifiers such as, for example, lymphokines, interleukin-I (IL-1), interleukin-2 (IL-2), interleukin-6 (IL-6), granulocyte macrophage colony stimulating factor (GMCSF), granulocyte colony stimulating factor (G-CSF), erythropoietin (EPO), pro-coagulants such as tissue factor and tissue factor variants, pro-apoptotic agents such FAS-ligand, fibroblast growth factors (FGF), nerve growth factor and other growth factors. Each must be in a form that can enter a cell or otherwise exert a therapeutic effect when in the vicinity thereof.
- Thus, therapeutic agents, include, but are not limited to, anti-tumor, anti-angiogenic, pro-apoptotic, anti-cancer and anti-mitotic agents. These are conjugated, optionally via a linker, to a substrate, such as peptidic substrate, which is a substrate for the protease.
- Among the therapeutic agents are cytotoxic agents that include, in general, but are not limited to, alkylating agents, toxins, antiproliferative agents and tubulin binding agents. Classes of cytotoxic agents for use herein include, for example, the anthracycline family of drugs, the vinca drugs, the mitomycins, the bleomycins, the cytotoxic nucleosides, the pteridine family of drugs, diynenes, the maytansinoids, the epothilones, the taxanes and the podophyllotoxins.
- Exemplary members of those classes include, for example, doxorubicin, carminomycin, daunorubicin, aminopterin, methotrexate, methopterin, dichloro-methotrexate, mitomycin C, porfiromycin, 5-fluorouracil, 6-mercaptopurine, cytosine arabinoside, podophyllotoxin, or podophyllotoxin derivatives such as etoposide or etoposide phosphate, melphalan, vinblastine, vincristine, leurosidine, vindesine, leurosine, maytansinol, epothilone A or B, taxotere, taxol and the like. Other such therapeutic agents include estramustine, cisplatin, combretastatin and analogs, and cyclophosphamide. One skilled in the art can make chemical modifications to the desired therapeutic agent in order to make reactions of that compound more convenient for purposes of preparing the conjugates.
- Particular therapeutic agents include the following drugs. One skilled in the art understands that these structural formulae are exemplary only and that such compounds or derivatives or analogs thereof have acquired in the art different generic or trivial names.
-
- in which
- R12 is amino or hydroxy;
- R7 is hydrogen or methyl;
- R8 is hydrogen, fluoro, chloro, bromo or iodo;
- R9 is hydroxy or a moiety which completes a salt of the carboxylic acid.
-
- in which R10 is hydrogen or methyl.
-
- in which R11 is hydroxy, amino, C1-C3 alkylamino, di(C1-C3 alkyl)amino, C4-C6 polymethylene amino, —NHCH2CH2CH2CH2NH—C(NH)NH2 or —NHCH2CH2CH2S+(CH3)2.
-
-
-
-
- in which
- R13 is hydrogen or methyl; and
- R14 is methyl or thienyl or a phosphate salt thereof.
-
- in which
- when R17 and R18 are taken singly, R15 is H, CH3 or CHO; and
- R18 is H, and one of R16 and R17 is ethyl and the other is H or OH;
- when R17 and R18 are taken together with the carbons to which they are attached, they form an oxirane ring in which case R16 is ethyl; and
- R19 is hydrogen, (C1-C3 alkyl)-CO, or chlorosubstituted (C1-C3 alkyl)-CO.
-
- where the peptidic substrate is as described above for formulae I and II; L is a linker such as —NH—(CH2)u—T—(CH2)u—NH—; Xn is
- a) hydrogen,
- b) —(C═O)R1a,
-
- d)
- e)
- f) ethoxysquarate; and
- g) cotininyl;
- R1 and R2 are independently hydrogen, OH, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 aralkyl and aryl;
- R1a is C1-C6-alkyl, hydroxylated C3-C8-cycloalkyl, polyhydroxylated C3-C8-cycloalkyl, hydroxylated aryl, polyhydroxylated aryl or aryl,
- R19 is hydrogen, (C1-C3 alkyl)-CO, or chlorosubstituted (C1-C3 alkyl)-CO;
- T is selected from cyclopentyl, cyclohexyl, cycloheptyl or bicyclo[2.2.2]octanyl;
- a is 1, 2, 3 or 4;
- b is zero or an integer between 1 and 100;
- c is 0 or 1, provided that if b is zero, c is 1;
- g is 1, 2 or 3;
- u is 0, 1, 2 or 3;
- or a pharmaceutically acceptable derivative thereof.
-
-
- R22 is hydrogen, methyl, bromo, fluoro, chloro or iodo;
- R23 is —OH or —NH2;
- R24 is hydrogen, bromo, chloro or lodo.
-
-
-
- in which
- Ra is —CH3, —CH2OH, —CH2OCO(CH2)3CH3, or —CH2OCOCH(OC2H5)2;
- Rb is —OCH3, —OH or —H;
- Rc is —NH2, —NHCOCF3, 4-morpholinyl, 3-cyano-4-morpholinyl, 1-piperidinyl, 4-methoxy-1-piperidinyl, benzylamine, dibenzylamine, cyanomethylamine, or 1-cyano-2-methoxyethyl amine;
- R5 is —OH —OTHP or —H; and
- R6 is —OH or —H provided that R6 is not —OH when R5 is —OH or —OTHP.
- Table 2, which follows, provides a number of anthracycline drugs and their generic or trivial names:
Compound Ra Rb Rc R5 R6 daunorubicina CH3 OCH3 NH2 OH H doxorubicinb CH2OH OCH3 NH2 OH H detorubicin CH2OCOCH(OC2H5)2 OCH3 NH2 OH H carminomycin CH3 OH NH2 OH H idarubicin CH3 H NH2 OH H epirubicin CH2OH OCH3 NH2 OH OH esorubicin CH2OH OCH3 NH2 H H THP CH2OH OCH3 NH2 OTHP H AD-32 CH2OCO(CH2)3CH3 OCH3 NHCOCF3 OH H - In one embodiment, when the therapeutic agent is doxorubicin, it is conjugated to the peptidic substrate via the amino group of the aminoglycoside moiety of doxorubicin.
-
- where R is PhC(O) or t-BuOC(O).
- In one embodiment, when the therapeutic agent is taxol (R=C(O)Ph), the peptidic substrate is conjugated to the secondary hydroxyl group of the cyclohexane moiety of taxol.
- p. Ribosome-Inactivating Proteins
- Ribosome-inactivating proteins (RIPs), which include ricin, abrin and saporin, are plant proteins that catalytically inactivate eukaryotic ribosomes. RIPS inactivate ribosomes by interfering with the protein elongation step of protein synthesis. For example, the RIP saporin (hereinafter also referred to as SAP) has been shown to enzymatically inactivate 60S ribosomes by cleavage of the n-glycosidic bond of the adenine at position 4324 in the rat 28S ribosomal RNA (rRNA). Some RIPs, such as the toxins abrin and ricin, contain two constituent chains: a cell-binding chain that mediates binding to cell surface receptors and internalization of the molecule; and an enzymatically active chain responsible for protein synthesis inhibitory activity. Such RIPs are type II RIPs. Other RIPs, such as the saporins, are single chains and are designated type I RIPs. Because such RIPs lack a cell-binding chain, they are less toxic to whole cells than the RIPs that have two chains. Two chain RIPs are generally used for conjugation herein, unless a single chain is further conjugated to an agent, such as a growth factor that mediates binding and internalization.
- Several structurally related RIP's have been isolated from seeds and leaves of the plantSaponaria officinalis (soapwort). Among these, SAP-6 is the most active and abundant, representing 7% of total seed proteins. Saporin is very stable, has a high isoelectric point, does not contain carbohydrates, and is resistant to denaturing agents, such as sodium dodecyl sulfate (SDS), and a variety of proteases. The amino acid sequences of several saporin-6 isoforms from seeds are known and there appear to be families of saporin RIPs differing in a few amino acid residues. Because saporin is a type I RIP, it does not possess a cell-binding chain. Consequently, its toxicity to whole cells is much lower than the other toxins, such as ricin and abrin. When internalized by eukaryotic cells, however, its cytotoxicity is 100- to 1000-fold more potent than ricin A chain.
- 4. Exemplary Conjugates
- The conjugates provided herein, are prepared by identifying suitable peptidic substrates for the targeted cell surface protease, or a soluble, shed or released form thereof, and forming a conjugate of the peptidic substrate(s) with a therapeutic agent(s). Exemplary conjugates, containing peptidic substrates designed, for example, for cleavage by MTSP1, endotheliase 1 and urokinase, are described. It is understood that upon identification of a cell surface protease, including cell-associated and cell-localized proteases, or a soluble, shed or released form thereof, in or associated with a cell involved in a disease or other conditions of interest, or with a cell present in the vicinity of a cell or tissue involved in or associated with a disease or other condition of interest, suitable peptidic substrates therefor can be empirically designed and then conjugated to therapeutic agents as exemplified herein.
- In certain embodiments, the conjugates for use in the compositions and methods provided herein include:
- Ac-Leu-Arg-Ala-Quat-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 46);
- Ac-Leu-Arg-Ala-Quat-Ala-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 47);
- Ac-Leu-Arg-Ser-Quat-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 48);
- Ac-Leu-Arg-Ser-Quat-Ala-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 49);
- Ac-Leu-Arg-Pro-Arg-Phe-Lys-Ile-Ile-(therapeutic agent) (SEQ ID NO: 50);
- Ac-Arg-Pro-Arg-Phe-Lys-Ile-Ile-(therapeutic agent) (SEQ ID NO: 51);
- Ac-Pro-Arg-Phe-Lys-Ile-Ile-(therapeutic agent) (SEQ ID NO: 52);
- Ac-Leu-Arg-Ser-Lys-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 53);
- Ac-Arg-Ser-Lys-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 54);
- Ac-Ser-Lys-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 55);
- Ac-Leu-Arg-Pro-Arg-Phe-Arg-Ile-Ile-(therapeutic agent) (SEQ ID NO: 56);
- Ac-Arg-Pro-Arg-Phe-Arg-Ile-Ile-(therapeutic agent) (SEQ ID NO: 57);
- Ac-Pro-Arg-Phe-Arg-Ile-Ile-(therapeutic agent) (SEQ ID NO: 58);
- Ac-Leu-Arg-Ser-Arg-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 59);
- Ac-Arg-Ser-Arg-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 60); and
- Ac-Ser-Arg-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 61).
- In further embodiments herein, the conjugates are Ac-Leu-Arg-Ala-Quat-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 62); Ac-Leu-Arg-Ala-Quat-Ala-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 63); Ac-Leu-Arg-Ser-Quat-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 64); and Ac-Leu-Arg-Ser-Quat-Ala-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 65).
- In other embodiments herein, the conjugates are
- Ac-Leu-Arg-Pro-Arg-Phe-Lys-Ile-Ile-(therapeutic agent) (SEQ ID NO: 66);
- Ac-Arg-Pro-Arg-Phe-Lys-Ile-Ile-(therapeutic agent) (SEQ ID NO: 67);
- Ac-Pro-Arg-Phe-Lys-Ile-Ile-(therapeutic agent) (SEQ ID NO: 68);
- Ac-Leu-Arg-Ser-Lys-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 69);
- Ac-Arg-Ser-Lys-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 70);
- Ac-Ser-Lys-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 71);
- Ac-Leu-Arg-Pro-Arg-Phe-Arg-Ile-Ile-(therapeutic agent) (SEQ ID NO: 72);
- Ac-Arg-Pro-Arg-Phe-Arg-Ile-Ile-(therapeutic agent) (SEQ ID NO: 73);
- Ac-Pro-Arg-Phe-Arg-Ile-Ile-(therapeutic agent) (SEQ ID NO: 74);
- Ac-Leu-Arg-Ser-Arg-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 75);
- Ac-Arg-Ser-Arg-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 76); and
- Ac-Ser-Arg-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 77).
- In other embodiments, the conjugates for use herein include the following:
- pyroGlu-Pro-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 78);
- CH3SO2-D-HHT-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 79);
- N-p-tosyl-Gly-Pro-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 80);
- Benzoyl-Val-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 81);
- CH3SO2-D-HHT-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 82);
- N-α-Z-D-Arg-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 83) (Z=benzyloxycarbonyl);
- pyroGlu-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 84);
- H-D-Ile-Pro-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 85); Cbo-L-(γ)Glu(α-t-BuO)-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 86) (Cbo=carbobenzoxy);
- H-D-Pro-Phe-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 87);
- H-D-Val-Leu-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 88);
- Bz-Ile-Glu(γ-OH)-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 89) (Bz=benzoyl);
- Bz-Ile-Glu(γ-OMe)-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 90);
- Benzoyl-Pro-Phe-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 91);
- H-D-Phe-Pip-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 92);
- H-D-Val-Leu-Lys-Ala-Ala-(therapeutic agent) (SEQ ID NO: 93);
- H-D-Nle-HHT-Lys-Ala-Ala-(therapeutic agent) (SEQ ID NO: 94);
- Pyr-Arg-Thr-Lys-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 95);
- H-Arg-Gln-Arg-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 96);
- Boc-Gln-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 97);
- Z-Arg-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 98);
- H-D-HHT-Ala-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 99);
- H-D-CHT-Gly-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 100);
- MeSO2-dPhe-Pro-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 101);
- δ-Z-D-Lys-Pro-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 102); and
- CH3SO2-D-CHA-But-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 103).
- In another embodiment, the conjugates for use in the compositions and methods provided herein include:
- Ac-Arg-Gln-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 104);
- Ac-Arg-Arg-Gln-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 105);
- Ac-Leu-Arg-Arg-Gin-Ser-Arg-Ala-Ala-(therapeutic agent) (SEQ ID NO: 106);
- Ac-Arg-Gln-Ser-Arg-Ala-(therapeutic agent) (SEQ ID NO: 107);
- Ac-Arg-Arg-Gln-Ser-Arg-Ala-(therapeutic agent) (SEQ ID NO: 108);
- Ac-Leu-Arg-Arg-Gln-Ser-Arg-Gly-Gly-(therapeutic agent) (SEQ ID NO: 109);
- Ac-Leu-Arg-Arg-Gln-Ser-Arg-Ala-(therapeutic agent) (SEQ ID NO: 110);
- Ac-Arg-Arg-Gln-Ser-Arg-Ile-(therapeutic agent) (SEQ ID NO: 111); and
- Ac-Leu-Arg-Arg-Gln-Ser-Arg-Ala-Ile-(therapeutic agent) (SEQ ID NO: 112).
- In certain embodiments, the conjugates for use in the compositions and methods provided herein include:
- Ac-Leu-Arg-Ala-Quat-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 113);
- Ac-Leu-Arg-Ala-Quat-Ala-Arg-Ser-Leu-(therapeutic,agent) (SEQ ID NO: 114);
- Ac-Leu-Arg-Ser-Quat-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 115);
- Ac-Leu-Arg-Ser-Quat-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 116);
- Ac-Leu-Arg-Pro-Arg-Phe-Lys-Ser-Leu-(therapeutic agent) (SEQ ID NO: 117);
- Ac-Arg-Pro-Arg-Phe-Lys-Ser-Leu-(therapeutic agent) (SEQ ID NO: 118);
- Ac-Pro-Arg-Phe-Lys-Ser-Leu-(therapeutic agent) (SEQ ID NO: 119);
- Ac-Leu-Arg-Ser-Lys-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 120);
- Ac-Arg-Ser-Lys-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 121);
- Ac-Ser-Lys-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 122);
- Ac-Leu-Arg-Pro-Arg-Phe-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 123);
- Ac-Arg-Pro-Arg-Phe-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 124);
- Ac-Pro-Arg-Phe-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 125);
- Ac-Leu-Arg-Ser-Arg-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 126);
- Ac-Arg-Ser-Arg-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 127); and
- Ac-Ser-Arg-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 128).
- In further embodiments herein, the conjugates are Ac-Leu-Arg-Ala-Quat-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 129); Ac-Leu-Arg-Ala-Quat-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 130); Ac-Leu-Arg-Ser-Quat-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 131); and Ac-Leu-Arg-Ser-Quat-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 132).
- In other embodiments herein, the conjugates are
- Ac-Leu-Arg-Pro-Arg-Phe-Lys-Ser-Leu-(therapeutic agent) (SEQ ID NO: 133);
- Ac-Arg-Pro-Arg-Phe-Lys-Ser-Leu-(therapeutic agent) (SEQ ID NO: 134);
- Ac-Pro-Arg-Phe-Lys-Ser-Leu-(therapeutic agent) (SEQ ID NO: 135);
- Ac-Leu-Arg-Ser-Lys-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 136);
- Ac-Arg-Ser-Lys-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 137);
- Ac-Ser-Lys-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 138);
- Ac-Leu-Arg-Pro-Arg-Phe-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 139);
- Ac-Arg-Pro-Arg-Phe-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 140);
- Ac-Pro-Arg-Phe-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 141);
- Ac-Leu-Arg-Ser-Arg-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 142);
- Ac-Arg-Ser-Arg-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 143); and
- Ac-Ser-Arg-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 144).
- In other embodiments, the conjugates for use herein include the following:
- pyroGlu-Pro-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 145); CH3SO2-D-HHT-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 146);
- N-p-tosyl-Gly-Pro-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 147);
- Benzoyl-Val-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 148);
- CH3SO2-D-HHT-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 149);
- N-α-Z-D-Arg-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 150) (Z=benzyloxycarbonyl);
- pyroGlu-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 151);
- H-D-Ile-Pro-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 152);
- Cbo-L-(γ)Glu(α-t-BuO)-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 153) (Cbo=carbobenzoxy);
- H-D-Pro-Phe-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 154);
- H-D-Val-Leu-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 155);
- Bz-Ile-Glu(γ-OH)-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 156) (Bz=benzoyl);
- Bz-Ile-Glu(γ-OMe)-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 157);
- Benzoyl-Pro-Phe-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 158);
- H-D-Phe-Pip-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 159);
- H-D-Val-Leu-Lys-Ser-Leu-(therapeutic agent) (SEQ ID NO: 160);
- H-D-Nle-HHT-Lys-Ser-Leu-(therapeutic agent) (SEQ ID NO: 161);
- Pyr-Arg-Thr-Lys-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 162);
- H-Arg-Gln-Arg-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 163);
- Boc-Gln-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 164);
- Z-Arg-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 165);
- H-D-HHT-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 166);
- H-D-CHT-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 167);
- MeSO2-dPhe-Pro-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 168);
- δ-Z-D-Lys-Pro-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 169); and
- CH3SO2-D-CHA-But-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 170).
- In another embodiment, the conjugates for use in the compositions and methods provided herein include:
- Ac-Arg-Gln-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 171);
- Ac-Arg-Arg-Gln-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 172);
- Ac-Leu-Arg-Arg-Gln-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 173);
- Ac-Arg-Gln-Ser-Arg-Leu-(therapeutic agent) (SEQ ID NO: 174);
- Ac-Arg-Arg-Gln-Ser-Arg-Leu-(therapeutic agent) (SEQ ID NO: 175);
- Ac-Leu-Arg-Arg-Gln-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 176);
- Ac-Leu-Arg-Arg-Gln-Ser-Arg-Leu-(therapeutic agent) (SEQ ID NO: 177);
- Ac-Arg-Arg-Gln-Ser-Arg-Leu-(therapeutic agent) (SEQ ID NO: 178); and
- Ac-Leu-Arg-Arg-Gln-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 179).
- In other embodiments, the conjugates provided herein include:
- Ac-Arg-Gln-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 180);
- Ac-Arg-Gln-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 181);
- Ac-Arg-Gln-Phe-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 182);
- Ac-Arg-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 183);
- Ac-Arg-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 184);
- Ac-Arg-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 185);
- Ac-Arg-Phe-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 186);
- Ac-Gln-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 187);
- Ac-Gln-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 188);
- Ac-Gln-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 189); and
- Ac-Gln-Phe-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 190).
- In further embodiments, the conjugates for use in the compositions and methods provided herein include:
- Ac-Leu-Arg-Ala-Quat-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 191);
- Ac-Leu-Arg-Ala-Quat-Ala-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 192);
- Ac-Leu-Arg-Ser-Quat-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 193);
- Ac-Leu-Arg-Ser-Quat-Ala-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 194);
- Ac-Leu-Arg-Pro-Arg-Phe-Lys-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 195);
- Ac-Arg-Pro-Arg-Phe-Lys-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 196);
- Ac-Pro-Arg-Phe-Lys-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 197);
- Ac-Leu-Arg-Ser-Lys-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 198);
- Ac-Arg-Ser-Lys-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 199);
- Ac-Ser-Lys-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 200);
- Ac-Leu-Arg-Pro-Arg-Phe-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 201);
- Ac-Arg-Pro-Arg-Phe-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 202);
- Ac-Pro-Arg-Phe-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 203);
- Ac-Leu-Arg-Ser-Arg-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 204);
- Ac-Arg-Ser-Arg-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 205); and
- Ac-Ser-Arg-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 206).
- In further embodiments herein, the conjugates are Ac-Leu-Arg-Ala-Quat-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 207); Ac-Leu-Arg-Ala-Quat-Ala-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 208); Ac-Leu-Arg-Ser-Quat-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 209); and Ac-Leu-Arg-Ser-Quat-Ala-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 210).
- In other embodiments herein, the conjugates are
- Ac-Leu-Arg-Pro-Arg-Phe-Lys-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 211);
- Ac-Arg-Pro-Arg-Phe-Lys-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 212);
- Ac-Pro-Arg-Phe-Lys-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 213);
- Ac-Leu-Arg-Ser-Lys-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 214);
- Ac-Arg-Ser-Lys-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 215);
- Ac-Ser-Lys-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 216);
- Ac-Leu-Arg-Pro-Arg-Phe-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 217);
- Ac-Arg-Pro-Arg-Phe-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 218);
- Ac-Pro-Arg-Phe-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 219);
- Ac-Leu-Arg-Ser-Arg-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 220);
- Ac-Arg-Ser-Arg-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 221); and
- Ac-Ser-Arg-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 222).
- In other embodiments, the conjugates for use herein include the following:
- pyroGlu-Pro-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 223);
- CH3SO2-D-HHT-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 224);
- N-p-tosyl-Gly-Pro-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 225); Benzoyl-Val-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 226); CH3SO2-D-HHT-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 227);
- N-α-Z-D-Arg-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 228) (Z=benzyloxycarbonyl);
- pyroGlu-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 229);
- H-D-Ile-Pro-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 230);
- Cbo-L-(γ)Glu(α-t-BuO)-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 231) (Cbo=carbobenzoxy);
- H-D-Pro-Phe-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 232);
- H-D-Val-Leu-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 233);
- Bz-Ile-Glu(γ-OH)-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 234) (Bz=benzoyl);
- Bz-Ile-Glu(γ-OMe)-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 235);
- Benzoyl-Pro-Phe-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 236);
- H-D-Phe-Pip-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 237);
- H-D-Val-Leu-Lys-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 238);
- H-D-Nle-HHT-Lys-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 239);
- Pyr-Arg-Thr-Lys-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 240);
- H-Arg-Gln-Arg-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 241);
- Boc-Gln-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 242);
- Z-Arg-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 243);
- H-D-HHT-Ala-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 244);
- H-D-CHT-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 245);
- MeSO2-dPhe-Pro-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 246);
- δ-Z-D-Lys-Pro-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 247); and
- CH3SO2-D-CHA-But-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 248).
- In another embodiment, the conjugates for use in the compositions and methods provided herein include:
- Ac-Arg-Gln-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 249);
- Ac-Arg-Arg-Gln-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 250);
- Ac-Leu-Arg-Arg-Gln-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 251);
- Ac-Arg-Gln-Ser-Arg-Leu-(therapeutic agent) (SEQ ID NO: 252);
- Ac-Arg-Arg-Gln-Ser-Arg-Leu-(therapeutic agent) (SEQ ID NO: 253);
- Ac-Leu-Arg-Arg-Gln-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 254);
- Ac-Leu-Arg-Arg-Gln-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 255);
- Ac-Arg-Arg-Gln-Ser-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 256); and
- Ac-Leu-Arg-Arg-Gln-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 257).
- In other embodiments, the conjugates provided herein include:
- Ac-Arg-Gln-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 258);
- Ac-Arg-Gln-Ala-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 259);
- Ac-Arg-Gln-Phe-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 260);
- Ac-Arg-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 261);
- Ac-Arg-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 262);
- Ac-Arg-Ala-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 263);
- Ac-Arg-Phe-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 264);
- Ac-Gln-Ser-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 265);
- Ac-Gln-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 266);
- Ac-Gln-Ala-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 267); and
- Ac-Gln-Phe-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 268).
- In another embodiment, the conjugates provided herein include:
- Ac-Gly-dSer-Ala-Arg-Ser-Ala-(therapeutic agent) (SEQ ID NO: 569);
- Ac-Arg-Gly-dSer-Ala-Arg-Ser-Ala-(therapeutic agent) (SEQ ID NO: 570);
- Ac-Gly-Ser-Gly-Arg-Ser-Ala-(therapetutic agent) (SEQ ID NO: 571);
- Ac-Arg-Gly-Ser-Gly-Arg-Ser-Ala-(therapetutic agent) (SEQ ID NO: 572);
- Ac-Leu-Arg-Gly-Ser-Gly-Arg-Ser-Ala-(therapetutic agent) (SEQ ID NO: 573);
- Ac-Leu-Arg-Gly-dSer-Ala-Arg-Ser-Ala-(therapetutic agent) (SEQ ID NO: 574);
- Ac-Cys(Me)-Pro-Gly-Arg-Val-Val-(therapeutic agent) (SEQ ID NO: 575);
- Ac-Arg-Cys(Me)-Pro-Gly-Arg-Val-Val-(therapeutic agent) (SEQ ID NO: 577);
- Ac-Arg-Arg-Cys(Me)-Pro-Gly-Arg-Val-Val-(therapeutic agent) (SEQ ID NO: 578);
- Ac-Val-Ser-Ala-Arg-Met-Ala-(therapeutic agent) (SEQ ID NO: 579);
- Ac-Ile-Val-Ser-Ala-Arg-Met-Ala-(therapeutic agent) (SEQ ID NO: 580);
- Ac-Val-Ile-Val-Ser-Ala-Arg-Met-Ala-(therapeutic agent) (SEQ ID NO: 581);
- Ac-Val-Ile-Val-Ser-Ala-Arg-nLeu-Ala-(therapeutic agent) (SEQ ID NO: 582);
- Ac-Val-Ser-Ala-Arg-nLeu-Ala-(therapeutic agent) (SEQ ID NO: 583);
- Ac-Ile-Val-Ser-Ala-Arg-nLeu-Ala-(therapeutic agent) (SEQ ID NO: 584);
- Ac-Gly-Ser-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 585);
- Ac-Gly-Ser-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 586);
- Ac-Gly-Ser-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 587);
- Ac-Ser-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 588);
- Ac-Ser-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 589);
- Ac-Ser-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 590);
- Ac-Arg-Gly-Ser-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 591);
- Ac-Arg-Gly-Ser-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 592);
- Ac-Arg-Gly-Ser-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 593);
- Ac-Leu-Arg-Gly-Ser-Gly-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 594);
- Ac-Leu-Arg-Gly-Ser-Gly-Arg-Ser-Ser-Leu-(therapeutic agent) (SEQ ID NO: 595); and
- Ac-Leu-Arg-Gly-Ser-Ala-Arg-Ser-Leu-(therapeutic agent) (SEQ ID NO: 596).
- In another embodiment, the conjugates provided herein are selected from:
- Ac-R-Q-G-R-S-L-(therapeutic agent) (SEQ ID NO: 491);
- Ac-R-Q-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 492);
- Ac-R-Q-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 493);
- Ac-R-Q-G-R-S-nV-(therapeutic agent) (SEQ ID NO: 494);
- Ac-R-Q-G-R-S-F-(therapeutic agent) (SEQ ID NO: 495);
- Ac-R-Q-G-R-A-L-(therapeutic agent) (SEQ ID NO: 496);
- Ac-R-Q-G-R-A-L-(therapeutic agent) (SEQ ID NO: 497);
- Ac-R-Q-G-R-A-nL-(therapeutic agent) (SEQ ID NO: 498);
- Ac-R-Q-G-R-A-nL-(therapeutic agent) (SEQ ID NO: 499);
- Ac-R-Q-G-R-A-nV-(therapeutic agent) (SEQ ID NO: 500);
- Ac-R-Q-G-R-A-Cha-(therapeutic agent) (SEQ ID NO: 501);
- Ac-R-Q-G-R-A-F-(therapeutic agent) (SEQ ID NO: 502);
- Ac-R-N-G-R-S-L-(therapeutic agent) (SEQ ID NO: 503);
- Ac-R-N-G-R-A-nL-(therapeutic agent) (SEQ ID NO: 504);
- Ac-R-Q-A-R-S-L-(therapeutic agent) (SEQ ID NO: 505);
- Ac-R-Q-A-R-S-nL-(therapeutic agent) (SEQ ID NO: 506);
- Ac-R-Q-A-R-S-nV-(therapeutic agent) (SEQ ID NO: 507);
- Ac-R-Q-A-A-S-Cha-(therapeutic agent) (SEQ ID NO: 508);
- Ac-R-Q-A-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 509);
- Ac-R-Q-A-R-T-nL-(therapeutic agent) (SEQ ID NO: 510);
- Ac-R-Q-A-R-A-L-(therapeutic agent) (SEQ ID NO: 511);
- Ac-R-Q-A-R-A-nL-(therapeutic agent) (SEQ ID NO: 512);
- Ac-R-Q-A-R-A-nV-(therapeutic agent) (SEQ ID NO: 513);
- Ac-R-Q-A-R-A-Cha-(therapeutic agent) (SEQ ID NO: 514);
- Ac-R-Q-S-R-A-A-(therapeutic agent) (SEQ ID NO: 515);
- Ac-R-Q-S-R-A-(therapeutic agent) (SEQ ID NO: 516);
- Ac-R-Q-S-R-A-nL-(therapeutic agent) (SEQ ID NO: 517);
- Ac-R-Q-S-R-A-L-(therapeutic agent) (SEQ ID NO: 518);
- Ac-R-Q-S-R-A-nV-(therapeutic agent) (SEQ ID NO: 519);
- Ac-R-Q-S-R-A-Cha-(therapeutic agent) (SEQ ID NO: 520);
- Ac-R-Q-S-R-S-S-L-(therapeutic agent) (SEQ ID NO: 521);
- Ac-R-Q-S-R-S-L-(therapeutic agent) (SEQ ID NO: 522);
- Ac-R-Q-S-R-S-nL-(therapeutic agent) (SEQ ID NO: 523);
- Ac-R-Q-S-R-S-nL-(therapeutic agent) (SEQ ID NO: 524);
- Ac-R-Q-S-R-S-nV-(therapeutic agent) (SEQ ID NO: 525);
- Ac-R-Q-S-R-S-allylG-(therapeutic agent) (SEQ ID NO: 526);
- Ac-R-Q-S-R-S-Cha-(therapeutic agent) (SEQ ID NO: 527);
- Ac-R-Q-S-R-T-nL-(therapeutic agent) (SEQ ID NO: 528);
- Ac-R-Q-T-R-S-S-L-(therapeutic agent) (SEQ ID NO: 529);
- Ac-R-Q-T-R-S-L-(therapeutic agent) (SEQ ID NO: 530);
- Ac-R-N-S-R-S-nL-(therapeutic agent) (SEQ ID NO: 531);
- Ac-R-Q-F-R-S-L-(therapeutic agent) (SEQ ID NO: 532);
- Ac-R-Q-F-R-S-nL-(therapeutic agent) (SEQ ID NO: 534);
- Ac-R-Q-F-R-S-nV-(therapeutic agent) (SEQ ID NO: 535);
- Ac-R-Q-F-R-S-nL-(therapeutic agent) (SEQ ID NO: 536);
- Ac-R-Q-F-R-S-Cha-(therapeutic agent) (SEQ ID NO: 537);
- Ac-R-Q-F-R-A-L-(therapeutic agent) (SEQ ID NO: 538);
- Ac-R-Q-F-R-A-nL-(therapeutic agent) (SEQ ID NO: 539);
- Ac-R-Q-F-R-A-nV-(therapeutic agent) (SEQ ID NO: 540);
- Ac-R-Q-F-R-A-Cha-(therapeutic agent) (SEQ ID NO: 541);
- Ac-Q-S-R-S-S-nL-(therapeutic agent) (SEQ ID NO: 542);
- MeOCO-Quat2-G-R-S-L-(therapeutic agent) (SEQ ID NO: 483);
- MeOCO-Quat3-G-R-S-L-(therapeutic agent) (SEQ ID NO: 484);
- MeOCO-Quat-G-R-S-L-(therapeutic agent) (SEQ ID NO: 485);
- MeOCO-Quat4-G-R-S-L-(therapeutic agent) (SEQ ID NO: 486);
- MeOCO-Quat5-G-R-S-L-(therapeutic agent) (SEQ ID NO: 487);
- MeOCO-Quat2-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 488);
- MeOCO-Quat4-G-R-S-L-(therapeutic agent) (SEQ ID NO: 489);
- MeOCO-Quat2-G-R-S-L-(therapeutic agent) (SEQ ID NO: 490);
- Ac-Q-G-R-S-L-(therapeutic agent) (SEQ ID NO: 445);
- Ac-Q-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 446);
- Ac-Q-G-R-A-S-L-(therapeutic agent) (SEQ ID NO: 447);
- Ac-N-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 448);
- Ac-Q-G-R-S-S-nL-(therapeutic agent) (SEQ ID NO: 449);
- Ac-Q-G-R-S-S-nV-(therapeutic agent) (SEQ ID NO: 450);
- Ac-Q-G-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 451);
- Ac-Q-G-R-S-S-allylG-(therapeutic agent) (SEQ ID NO: 452);
- Ac-Q-G-R-S-S-allylG-(therapeutic agent) (SEQ ID NO: 453);
- Ac-Q-A-R-S-L-(therapeutic agent) (SEQ ID NO: 454);
- Ac-Q-A-R-S-S-L-(therapeutic agent) (SEQ ID NO: 455);
- Ac-Q-S-R-S-L-(therapeutic agent) (SEQ ID NO: 456);
- Ac-Q-S-R-S-S-nV-(therapeutic agent) (SEQ ID NO: 457);
- Ac-Q-S-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 458);
- Ac-Q-S-R-S-S-L-(therapeutic agent) (SEQ ID NO: 459);
- Ac-Q-T-R-S-S-L-(therapeutic agent) (SEQ ID NO: 460);
- Ac-Q-Aib-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 461);
- Ac-Q-Aib-R-S-S-L-(therapeutic agent) (SEQ ID NO: 462);
- Ac-Q-Abu-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 463);
- Ac-Q-Abu-R-S-S-L-(therapeutic agent) (SEQ ID NO: 464);
- Ac-Q-Cha-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 465);
- Ac-Q-F-R-S-L-(therapeutic agent) (SEQ ID NO: 466);
- Ac-Q-F-R-S-S-L-(therapeutic agent) (SEQ ID NO: 467);
- Ac-Q-Y-R-S-S-L-(therapeutic agent) (SEQ ID NO: 468);
- Ac-R-G-R-S-L-(therapeutic agent) (SEQ ID NO: 469);
- Ac-R-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 470);
- Ac-R-G-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 471);
- Ac-R-G-R-S-Cha-(therapeutic agent) (SEQ ID NO: 472);
- Ac-R-A-R-S-L-(therapeutic agent) (SEQ ID NO: 473);
- Ac-R-A-R-S-S-L-(therapeutic agent) (SEQ ID NO: 474);
- Ac-R-S-R-S-L-(therapeutic agent) (SEQ ID NO: 475);
- Ac-R-S-R-S-S-L-(therapeutic agent) (SEQ ID NO: 476);
- Ac-R-S-R-S-Cha-(therapeutic agent) (SEQ ID NO: 477);
- Ac-R-S-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 478);
- Ac-R-F-R-S-L-(therapeutic agent) (SEQ ID NO: 479);
- Ac-R-F-R-S-Cha-(therapeutic agent) (SEQ ID NO: 480);
- Ac-Y-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 481);
- Ac-M(O2)-S-R-S-L-(therapeutic agent) (SEQ ID NO: 482);
- Ac-R-R-Q-S-R-A-A-(therapeutic agent) (SEQ ID NO: 105);
- Ac-R-R-Q-S-R-I-(therapeutic agent) (SEQ ID NO: 610);
- Ac-R-R-Q-S-R-S-S-L-(therapeutic agent) (SEQ ID NO: 543);
- Ac-R-R-Q-S-R-S-L-(therapeutic agent) (SEQ ID NO: 544);
- Ac-R-G-S-G-R-S-L-(therapeutic agent) (SEQ ID NO: 545);
- Ac-R-G-S-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 546);
- Ac-R-G-S-G-R-A-nL-(therapeutic agent) (SEQ ID NO: 547);
- Ac-R-G-S-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 548);
- Ac-I-V-S-G-R-A-S-L-(therapeutic agent) (SEQ ID NO: 549);
- Ac-R-R-Q-S-R-A-(therapeutic agent) (SEQ ID NO: 108);
- Ac-R-R-Q-S-R-I-(therapeutic agent) (SEQ ID NO: 111);
- Ac-L-R-R-Q-S-R-A-A-(therapeutic agent) (SEQ ID NO: 106);
- Ac-L-R-R-Q-S-R-G-G-(therapeutic agent) (SEQ ID NO: 109);
- Ac-L-R-R-Q-S-R-A-(therapeutic agent) (SEQ ID NO: 110);
- Ac-L-R-R-Q-S-R-A-I-(therapeutic agent) (SEQ ID NO: 112);
- Ac-L-R-R-Q-S-R-A-I-(therapeutic agent) (SEQ ID NO: 611);
- Ac-L-R-R-Q-S-R-S-S-L-(therapeutic agent) (SEQ ID NO: 550); and
- Ac-L-R-R-Q-S-R-S-L-(therapeutic agent) (SEQ ID NO: 551);
- In another embodiment, the conjugates provided herein are selected from:
- Ac-S-G-R-S-L-(therapeutic agent) (SEQ ID NO: 362);
- Ac-S-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 363);
- Ac-S-G-R-S-S-S-L-(therapeutic agent) (SEQ ID NO: 364);
- Ac-S-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 365);
- Ac-S-G-R-S-nV-(therapeutic agent) (SEQ ID NO: 366);
isomer 1 - Ac-S-G-R-S-nV-(therapeutic agent) (SEQ ID NO: 367);
isomer 2 - Ac-S-G-R-S-G(hex)-(therapeutic agent) (SEQ ID NO: 368);
- Ac-S-G-R-S-Cha-(therapeutic agent) (SEQ ID NO: 369);
- Ac-S-G-R-S-hCha-(therapeutic agent) (SEQ ID NO: 370);
- Ac-S-A-R-S-L-(therapeutic agent) (SEQ ID NO: 371);
- Ac-S-A-R-S-S-L-(therapeutic agent) (SEQ ID NO: 372);
- Ac-S-S-R-S-nL-(therapeutic agent) (SEQ ID NO: 373);
- Ac-T-G-R-S-Abu-(therapeutic agent) (SEQ ID NO: 374);
- Ac-T-G-R-S-L-(therapeutic agent) (SEQ ID NO: 375);
- Ac-T-G-R-S-nV-(therapeutic agent) (SEQ ID NO: 376);
- Ac-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 377);
- Ac-T-G-R-S-G(hex)-(therapeutic agent) (SEQ ID NO: 378);
- Ac-T-G-R-S-Cha-(therapeutic agent) (SEQ ID NO: 379);
- Ac-T-G-R-S-hCha-(therapeutic agent) (SEQ ID NO: 380);
- Ac-T-G-R-T-Abu-(therapeutic agent) (SEQ ID NO: 381);
- Ac-T-G-R-hS-nL-(therapeutic agent) (SEQ ID NO: 382);
- Ac-T-G-R-Abu-nL-(therapeutic agent) (SEQ ID NO: 383);
- Ac-T-G-R-Abu-nV-(therapeutic agent) (SEQ ID NO: 384);
- Ac-T-G-F(Gn)-S-nL-(therapeutic agent) (SEQ ID NO: 385);
- Ac-T-G-F(Gn)-S-Cha-(therapeutic agent) (SEQ ID NO: 386);
- Ac-T-G-F(Gn)-Abu-nV-(therapeutic agent) (SEQ ID NO: 387);
- Ac-T-G-K(alloc)-S-nL-(therapeutic agent) (SEQ ID NO: 388);
- Ac-T-G-K-S-nL-(therapeutic agent) (SEQ ID NO: 389);
- Ac-T-G-hR-S-nL-(therapeutic agent) (SEQ ID NO: 390);
- Ac-(hS)G-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 391);
- MeOCO-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 392);
- PhSO2-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 393);
- MeOEtCO-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 394);
- MeO(EtO)2Ac-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 395);
- 4-oxo-Pentanoyl-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 396);
- 3,4-MethyidioxyPhAc-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 397);
- 2-PyridylAc-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 398);
- PhOAc-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 399);
- L-3-PhLactyl-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 400);
- MeOAc-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 401);
- PhAc-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 402);
- MeOEtOCO-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 403);
- MeOEtOAc-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 404);
- HOOCButa-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 405);
- Z-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 406);
- EtOCO-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 407);
- βA-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 408);
- Pent-4-ynoyl-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 409);
- NapAc-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 410);
- iBoc-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 411);
- HOAc-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 412);
- MeSucc-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 413);
- N,N-diMeGly-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 414);
- Succ-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 415);
- HCO-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 416);
- Ac-T-A-R-S-nL-(therapeutic agent) (SEQ ID NO: 417);
- Ac-T-A-F(Gn)-S-nL-(therapeutic agent) (SEQ ID NO: 418);
- Ac-T-A-R-Abu-nV-(therapeutic agent) (SEQ ID NO: 419);
- Ac-T-A-R-S-Abu-(therapeutic agent) (SEQ ID NO: 420);
- Ac-T-A-R-T-Abu-(therapeutic agent) (SEQ ID NO: 421);
- Ac-T-S(O-Me)-R-S-nL-(therapeutic agent) (SEQ ID NO: 422);
- Ac-T-hS-R-S-nL-(therapeutic agent) (SEQ ID NO: 423);
- Ac-T-(1-Me)H-R-S-nL-(therapeutic agent) (SEQ ID NO: 424);
- Ac-T-(3-Me)H-R-S-nL-(therapeutic agent) (SEQ ID NO: 425);
- Ac-T-H-R-S-nL-(therapeutic agent) (SEQ ID NO: 426);
- Ac-T-Sar-R-S-nL-(therapeutic agent) (SEQ ID NO: 427);
- Ac-T-nV-R-S-nL-(therapeutic agent) (SEQ ID NO: 428);
- Ac-T-nL-R-S-nL-(therapeutic agent) (SEQ ID NO: 429);
- Ac-T-A-R-S-Cha-(therapeutic agent) (SEQ ID NO: 430);
- Ac-T-Abu-R-S-nL-(therapeutic agent) (SEQ ID NO: 431);
- Ac-4,4diMeThr-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 432);
- Ac-hS-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 433);
- Ac-hS-G-R-hS-Cha-(therapeutic agent) (SEQ ID NO: 434);
- Ac-hS-G-R-S-Cha-(therapeutic agent) (SEQ ID NO: 435);
- Ac-hS-G-R-T-Cha-(therapeutic agent) (SEQ ID NO: 436);
- Ac-hS-A-R-S-Cha-(therapeutic agent) (SEQ ID NO: 437);
- Ac-N-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 438);
- Ac-Y-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 439);
- Ac-Y-G-R-S-Cha-(therapeutic agent) (SEQ ID NO: 440);
- Ac-Q-G-R-S-S-nL-(therapeutic agent) (SEQ ID NO: 441);
- Ac-Q-G-R-S-S-nV-(therapeutic agent) (SEQ ID NO: 442);
- Ac-L-R-G-S-G-R-S-A-(therapeutic agent) (SEQ ID NO: 573);
- Ac-L-R-G-S-G-R-S-L-(therapeutic agent) (SEQ ID NO: 342);
- Ac-L-R-G-S-G-R-S-L-(therapeutic agent) (SEQ ID NO: 343);
- Ac-L-R-G-S-G-R-S-S-nL-(therapeutic agent) (SEQ ID NO: 344);
- Ac-L-R-G-S-G-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 345);
- Ac-L-R-G-dS-A-R-S-A-(therapeutic agent) (SEQ ID NO: 574);
- Ac-L-R-G-S-A-R-S-S-L-(therapeutic agent) (SEQ ID NO: 346);
- Ac-L-R-G-S-A-R-S-L-(therapeutic agent) (SEQ ID NO: 347);
- Ac-L-R-G-S-A-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 348);
- Ac-L-R-G-S-A-R-S-S-nV-(therapeutic agent) (SEQ ID NO: 349);
- Ac-L-R-G-S-A-R-S-S-nL-(therapeutic agent) (SEQ ID NO: 350);
- Ac-V-I-V-S-G-R-A-L-(therapeutic agent) (SEQ ID NO: 351);
- Ac-V-I-V-S-A-R-S-L-(therapeutic agent) (SEQ ID NO: 352);
- Ac-V-I-V-S-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 353);
- Ac-V-I-V-S-A-R-M-A-(therapeutic agent) (SEQ ID NO: 354);
- Ac-V-I-V-S-A-R-nL-A-(therapeutic agent) (SEQ ID NO: 355);
- Ac-V-I-V-S-A-R-S-nL-(therapeutic agent) (SEQ ID NO: 356);
- Ac-V-I-V-S-A-R-S-Cha-(therapeutic agent) (SEQ ID NO: 357);
- Ac-V-I-V-S-A-R-S-Cha-(therapeutic agent) (SEQ ID NO: 358);
- Ac-V-I-V-S-A-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 359);
- Ac-R-R-(Me)C-P-G-R-V-V-(therapeutic agent) (SEQ ID NO: 360);
- Ac-R-R-nV-P-A-R-S-L-(therapeutic agent) (SEQ ID NO: 361);
- Ac-R-G-dS-A-R-S-A-(therapeutic agent) (SEQ ID NO: 309);
- Ac-R-G-S-G-R-S-A-(therapeutic agent) (SEQ ID NO: 310);
- Ac-R-G-S-G-R-A-L-(therapeutic agent) (SEQ ID NO: 311);
- Ac-R-G-S-G-R-S-L-(therapeutic agent) (SEQ ID NO: 312);
- Ac-R-G-S-G-R--S-nL-(therapeutic agent) (SEQ ID NO: 313);
- Ac-R-G-S-G-R-A-nL-(therapeutic agent) (SEQ ID NO: 314);
- Ac-R-G-S-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 315);
- Ac-R-G-S-G-R-S-Cha-(therapeutic agent) (SEQ ID NO: 316);
- Ac-R-G-S-G-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 317);
- Ac-R-G-S-A-R-S-Cha-(therapeutic agent) (SEQ ID NO: 318);
- Ac-R-G-S-A-R-S-S-(therapeutic agent) (SEQ ID NO: 319);
- Ac-R-G-S-A-R-S-nV-(therapeutic agent) (SEQ ID NO: 320);
- Ac-R-G-S-A-R-S-S-nV-(therapeutic agent) (SEQ ID NO: 321);
- Ac-R-G-S-A-R-S-L-(therapeutic agent) (SEQ ID NO: 322);
- Ac-R-(Me)C-P-G-R-V-V-(therapeutic agent) (SEQ ID NO: 323);
- Ac-R-(Me)C-P-G-R-V-V-(therapeutic agent) (SEQ ID NO: 324);
- Ac-R-C(Me)-P-G-R-S-L-(therapeutic agent) (SEQ ID NO: 325);
- Ac-R-L-P-G-R-S-L-(therapeutic agent) (SEQ ID NO: 326);
- Ac-R-V-P-G-R-S-L-(therapeutic agent) (SEQ ID NO: 327);
- Ac-R-V-P-G-R-S-L-(therapeutic agent) (SEQ ID NO: 328);
- Ac-R-nL-P-G-R-S-L-(therapeutic agent) (SEQ ID NO: 329);
- Ac-R-G(tBu)-P-A-R-S-L-(therapeutic agent) (SEQ ID NO: 330);
- Ac-R-L-P-A-R-S-L-(therapeutic agent) (SEQ ID NO: 331);
- Ac-R-V-P-A-R-S-L-(therapeutic agent) (SEQ ID NO: 332);
- Ac-R-nL-P-A-R-S-L-(therapeutic agent) (SEQ ID NO: 333);
- Ac-I-V-S-G-R-A-L-(therapeutic agent) (SEQ ID NO: 334);
- Ac-I-V-S-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 335);
- Ac-I-V-S-G-R-A-S-L-(therapeutic agent) (SEQ ID NO: 336);
- Ac-I-V-S-A-R-M-A-(therapeutic agent) (SEQ ID NO: 337);
- Ac-I-V-S-A-R-nL-A-(therapeutic agent) (SEQ ID NO: 338);
- Ac-I-V-S-A-R-S-L-(therapeutic agent) (SEQ ID NO: 339);
- Ac-I-V-S-A-R-S-nL-(therapeutic agent) (SEQ ID NO: 340);
- Ac-I-V-S-A-R-S-S-L-(therapeutic agent) (SEQ ID NO: 341);
- Ac-G-S-G-R-S-A-(therapeutic agent) (SEQ ID NO: 585);
- Ac-G-S-G-R-S-L-(therapeutic agent) (SEQ ID NO: 277);
- Ac-G-S-G-R-A-L-(therapeutic agent) (SEQ ID NO: 278);
- Ac-G-S-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 279);
- Ac-G-S-G-R-L-(therapeutic agent) (SEQ ID NO: 280);
- Ac-G-S-G-(4-guan)Phg-S-L-(therapeutic agent) (SEQ ID NO: 281);
- Ac-G-S-G-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 282);
- Ac-G-S-G-R-A-S-L-(therapeutic agent) (SEQ ID NO: 283);
- Ac-G-S-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 284);
- Ac-G-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 285);
- Succ-bA-T-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 286);
- Ac-G-T-G-R-S-hCha-(therapeutic agent) (SEQ ID NO: 287);
- Ac-G-hS-G-R-S-nL-(therapeutic agent) (SEQ ID NO: 288);
- Ac-G-dS-A-R-S-A-(therapeutic agent) (SEQ ID NO: 289);
- Ac-G-S-A-R-S-L-(therapeutic agent) (SEQ ID NO: 290);
- Ac-G-S-A-R-S-S-Cha-(therapeutic agent) (SEQ ID NO: 291);
- Ac-G-S-A-R-S-S-L-(therapeutic agent) (SEQ ID NO: 292);
- Ac-G-S-A-R-A-S-L-(therapeutic agent) (SEQ ID NO: 293);
- Ac-V-S-G-R-S-L-(therapeutic agent) (SEQ ID NO: 294);
- Ac-V-S-G-R-A-L-(therapeutic agent) (SEQ ID NO: 295);
- Ac-V-S-G-R-A-S-L-(therapeutic agent) (SEQ ID NO: 296);
- Ac-V-S-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 297);
- Ac-V-S-A-R-M-A-(therapeutic agent) (SEQ ID NO: 298);
- Ac-V-S-A-R-nL-A-(therapeutic agent) (SEQ ID NO: 299);
- Ac-V-S-A-R-S-nL-(therapeutic agent) (SEQ ID NO: 300);
- Ac-V-S-A-R-S-L-(therapeutic agent) (SEQ ID NO: 301);
- Ac-(Me)C-P-G-R-V-V-(therapeutic agent) (SEQ ID NO: 302);
- Ac-(Me)C-P-G-R-V-V-(therapeutic agent) (SEQ ID NO: 303);
- Ac-C(Me)-P-G-R-A-L-(therapeutic agent) (SEQ ID NO: 304);
- Ac-C(Me)-P-G-R-S-L-(therapeutic agent) (SEQ ID NO: 305);
- Ac-C(Me)-P-A-R-S-L-(therapeutic agent) (SEQ ID NO: 306);
- Ac-C(Me)-P-A-R-A-S-L-(therapeutic agent) (SEQ ID NO: 307);and
- Ac-G(tBu)-P-G-R-S-L-(therapeutic agent) (SEQ ID NO: 308);
- In another embodiment, the conjugates provided herein are selected from:
- Ac-Q-S-R-A-A-(therapeutic agent) (SEQ ID NO: 552);
- Ac-Q-S-R-S-A-(therapeutic agent) (SEQ ID NO: 553);
- Ac-Q-S-R-S-G-(therapeutic agent) (SEQ ID NO: 554);
- Ac-R-S-R-A-A-(therapeutic agent) (SEQ ID NO: 555);
- Ac-R-Q-S-R-A-A-(therapeutic agent) (SEQ ID NO: 556);
- Ac-R-Q-S-R-S-A-(therapeutic agent) (SEQ ID NO: 557); and
- Ac-R-Q-S-R-S-A-A-(therapeutic agent) (SEQ ID NO: 558);
- In another embodiment, the conjugates provided herein are selected from:
- Ac-R-G-S-G-R-S-A-(therapeutic agent) (SEQ ID NO: 559);
- Ac-S-G-R-A-A-(therapeutic agent) (SEQ ID NO: 560);
- Ac-S-G-R-S-A-(therapeutic agent) (SEQ ID NO: 561);
- Ac-S-G-R-S-S-A-(therapeutic agent) (SEQ ID NO: 562);
- Ac-S-G-R-A-S-A-(therapeutic agent) (SEQ ID NO: 563);
- Ac-S-G-R-S-G-(therapeutic agent) (SEQ ID NO: 564);
- Ac-S-G-R-S-S-G-(therapeutic agent) (SEQ ID NO: 565);
- Ac-S-G-R-S-G-A-(therapeutic agent) (SEQ ID NO: 566);
- Ac-S-G-R-S-G-G-(therapeutic agent) (SEQ ID NO: 567); and
- Ac-G-T-G-R-S-G-G-(therapeutic agent) (SEQ ID NO: 568);
- In another embodiment, the conjugates provided herein are selected from:
- Ac-L-R-R-Q-S-R-A-A-(therapeutic agent) (SEQ ID NO: 597);
- MeSO2-dA(Chx)-Abu-R-S-L-(therapeutic agent) (SEQ ID NO: 598);
- Ac-R-A-R-S-L-(therapeutic agent) (SEQ ID NO: 599);
- Ac-dA(Chx)-Abu-R-S-L-(therapeutic agent) (SEQ ID NO: 600);
- Ac-dA(Chx)-Abu-R-S-S-L-(therapeutic agent) (SEQ ID NO: 601);
- Ac-Q-G-R-S-S-L-(therapeutic agent) (SEQ ID NO: 602);
- MeOCO-dhF-P(OH)-R-S-S-L-(therapeutic agent) (SEQ ID NO: 603);
- MeOCO-Quat4-G-R-S-L-(therapeutic agent) (SEQ ID NO: 604);
- Ac-dCha-P(OH)-R-S-S-L-(therapeutic agent) (SEQ ID NO: 605);
- Ac-dCha-Abu-R-S-S-A-(therapeutic agent) (SEQ ID NO: 606);
- MeOCO-Quat2-G-R-S-L-(therapeutic agent) (SEQ ID NO: 607);
- MeOCO-Quat3-G-R-S-L-(therapeutic agent) (SEQ ID NO: 608); and
- MeOCO-Quat-G-R-S-L-(therapeutic agent) (SEQ ID NO: 609).
- It also understood that conjugates containing the above peptidic substrate portions can be prepared with other capping groups in place of Ac (see, e.g., the description herein of the capping groups Xn). Therapeutic agents for use in the above conjugates include, for example, cytotoxic agents, such as, but not limited to, a toxin such as abrin, ricin A, pseudomonas exotoxin shiga toxin, diphtheria toxin and other such toxins and toxic portions thereof; proteins such as tumor necrosis factor, interferons, such as α-interferon and gamma-interferon, pro-coagulants such as tissue factor and tissue factor variants, pro-apoptotic agents such FAS-ligand, nerve growth factor, platelet derived growth factor, tissue plasminogen activator; biological response modifiers such as, for example, lymphokines, interleukin-I (IL-I), interleukin-2 (IL-2), interleukin-6 (IL-6), granulocyte macrophage colony stimulating factor (GMCSF), granulocyte colony stimulating factor (G-CSF), fibroblast growth factors (FGFs) and other growth factors, the methotrexate group of drugs, the anthracycline family of drugs, the vinca alkaloid drugs, the mitomycins, the bleomycins, the cytotoxic nucleosides including cytosine arabinosides and difluoronucleosides, the pteridine family of drugs, diynenes, the taxanes and the podophyllotoxins. All such conjugates are within the scope of the instant disclosure and can be prepared and used as disclosed herein.
- Thus, the conjugates provided herein include, but are not limited to, those disclosed herein where the therapeutic agent is, e.g., doxorubicin, carminomycin, daunorubicin, detorubicin, idarubicin, epirubicin, esorubicin, THP, AD-32, aminopterin, methotrexate, methopterin, dichloromethotrexate, mitomycin C, porfiromycin, 5-fluorouracil, 6-mercaptopurine, cytosine arabinoside, podophyllotoxin, or podophyllotoxin derivatives such as etoposide or etoposide phosphate, melphalan, vinblastine, vincristine, leurosidine, vindesine, leurosine, taxol, estramustine, cisplatin, combretastatin and analogs, and cyclophosphamide. In one embodiment, the therapeutic agent is doxorubicin. In another embodiment, the therapeutic agent is taxol.
- Any conjugates corresponding to the above conjugates or any conjugates disclosed herein where the P1′ and/or P2′ residues are Ile in place of Ala are within the scope of the instant disclosure and can be prepared and used as disclosed herein.
- Any peptidic substrates formed by permutation and selection of amino acids from those set forth in the above definitions of P groups are contemplated.
- The peptidic substrates of the conjugates provided herein are synthesized from their constituent amino acids by conventional peptide synthesis techniques, such as by solid-phase technology. The peptidic substrates are then purified by reverse-phase high performance liquid chromatography (HPLC).
- The peptide acids can be prepared from their constituent Fmoc-aminoacids. Standard methods of peptide synthesis are disclosed, for example, in the following works: Synthesis Notes Section, NovaBiochem Catalog 2002/3, Schroeder et al., “The Peptides”, Vol. 1, Academic Press 1965; Bodansky et al., “Peptide Synthesis”, Interscience Publishers, 1966; McOmie (ed.) “Protective Groups in Organic Chemistry”, Plenum Press, 1973, Barany et al., “The Peptides: Analysis, Synthesis, Biology” 2,
Chapter 1, Academic Press, 1990, and Stewart et al., “Solid Phase Peptide Synthesis”, Second Edition, Pierce Chemical Company, 1994. The disclosures of these references are hereby incorporated by reference. - The pharmaceutically acceptable salts of the conjugates provided herein include the conventional non-toxic salts of the conjugates as formed, e.g., from non-toxic inorganic or organic acids. For example, such conventional non-toxic salts include those derived from inorganic acids such as hydrochloric, hydrobromic, sulfuric, sulfamic, phosphoric, nitric and the like: and the salts prepared from organic acids such as acetic, propionic, succinic, glycolic, stearic, lactic, malic, tartaric, citric, ascorbic, pamoic, maleic, hydroxymaleic, phenyl-acetic, glutamic, benzoic, salicylic, sulfanilic, 2-acetoxy-benzoic, fumaric, toluenesulfonic, methanesulfonic, ethane disulfonic, oxalic, isethionic, trifluoroacetic and the like.
- The conjugates provided herein that contain the peptidic moieties containing the cell surface protease cleavage site and a therapeutic agent can similarly be synthesized by techniques known to those of skill in the art. For example, a free amine moiety on the therapeutic agent can be covalently attached to the peptidic substrate at the carboxyl terminus such that an amide bond is formed. Similarly, an amide bond can be formed by covalently coupling an amine moiety of the peptidic substrate and a carboxyl moiety of the therapeutic agent. For these purposes a reagent such as 2-(1H-benzotriazol-1-yl)-1,3,3-tetramethyl-uronium hexafluorophosphate (known as HBTU) and 1-hyroxybenzotriazole hydrate (known as HOBT), dicyclohexyl-carbodiimide (DCC), N-ethyl-N-(3-dimethylaminopropyl)-carbodiimide (EDC), diphenyl-phosphorylazide (DPPA), benzotriazol-1-yl-oxy-tris-(dimethylamino)-phosphonium hexafluorophosphate (BOP) and the like, used in combination or singularly, can be utilized.
- Furthermore, the instant conjugates can be formed by a non-peptidyl bond between the cell surface protease cleavage site and a therapeutic agent. For example, the therapeutic agent can be covalently attached to the carboxyl terminus of the peptidic substrate via a hydroxyl moiety on the therapeutic agent, thereby forming an ester linkage. For this purpose a reagent such as a combination of HBTU and HOBT, a combination of BOP and imidazole, a combination of DCC and DMAP, and the like can be utilized. The carboxylic acid also can be activated by forming the nitro-phenyl ester or the like and reacted in the presence of DBU (1,8-diazabicyclo[5,4,O]undec-7-ene).
- The instant conjugates also can be formed by attachment of the peptidic substrate to the therapeutic agent via a linker unit. Such linker units include, for example, a biscarbonyl alkyl diradical whereby an amine moiety on the therapeutic agent is connected with the linker unit to form an amide bond and the amino terminus of the peptidic substrate is connected with the other end of the linker unit also forming an amide bond. Conversely, a diaminoalkyl diradical linker unit, whereby a carbonyl moiety on the cytotoxic agent is covalently attached to one of the amines of the linker unit while the other amine of the linker unit is covalently attached to the C-terminus of the peptidic substrate, also can be useful. Other such linker units which are stable to the physiological environment when not in the presence of a cell surface protease, or a soluble, shed or released form thereof, but are cleavable upon the cleavage of the cell surface protease proteolytic cleavage site, or a soluble, shed or released form thereof, are also envisioned. Furthermore, linker units can be utilized that, upon cleavage of the cell surface protease proteolytic cleavage site, remain attached to the therapeutic agent but do not significantly decrease the therapeutic activity of such a post-cleavage therapeutic agent derivative when compared with an unmodified therapeutic agent.
- One skilled in the art understands that in the synthesis of the conjugates provided herein, one can need to protect various reactive functionalities on the starting compounds and intermediates while a desired reaction is carried out on other portions of the molecule. After the desired reactions are complete, or at any desired time, normally such protecting groups will be removed by, for example, hydrolytic or hydrogenolytic means. Such protection and deprotection steps are conventional in organic chemistry. One skilled in the art is referred to Protective Groups in Organic Chemistry, McOmie, ed., Plenum Press, NY, N.Y. (1973); and, Protective Groups in Organic Synthesis, Greene, ed., John Wiley & Sons, NY, N.Y. (1991) for the teaching of protective groups which can be useful in the preparation of the conjugates provided herein.
- By way of example only, useful amino-protecting groups can include, for example, C1-C10 alkanoyl groups such as formyl, acetyl, dichloroacetyl, propionyl, hexanoyl, 3,3-diethylhexanoyl, γ-chlorobutryl, and the like; C1-C10 alkoxycarbonyl and C5-C15 aryloxycarbonyl groups such as tert-butoxycarbonyl, benzyloxycarbonyl, allyloxycarbonyl, 4-nitrobenzyloxycarbonyl, fluorenylmethyloxycarbonyl and cinnamoyloxy-carbonyl; halo(C1-C10)-alkoxycarbonyl such as 2,2,2-trichloroethoxy-carbonyl; and C1-C15 arylalkyl and alkenyl group such as benzyl, phenethyl, allyl, trityl, and the like. Other commonly used amino-protecting groups are those in the form of enamines prepared with β-keto-esters such as methyl or ethyl acetoacetate.
- Useful carboxy-protecting groups can include, for example, C1-C10 alkyl groups such as methyl, tert-butyl, decyl; halo C1-C10 alkyl such as 2,2,2-trichloroethyl, and 2-iodoethyl; C5-C15 arylalkyl such as benzyl, 4-methoxybenzy], 4-nitrobenzyl, triphenylmethyl, diphenyl-methyl; C1-C10 alkanoyloxymethyl such as acetoxy-methyl, propionoxymethyl and the like; and groups such as phenacyl, 4-halophenacyl, allyl, dimethylallyl, tri-(C1-C3 alkyl)silyl, such as trimethylsilyl, β-p-toluenesulfonylethyl, β-p-nitrophenyl-thioethyl, 2,4,6-trimethylbenzyl, β-methylthioethyl, phthalimidomethyl, 2,4-dinitro-phenylsulphenyl, 2-nitrobenzhydryl and related groups.
- Similarly, useful hydroxy protecting groups can include, for example, the formyl group, the chloroacetyl group, the benzyl group, the benzhydryl group, the trityl group, the 4-nitrobenzyl group, the trimethylsilyl group, the phenacyl group, the tert-butyl group, the methoxymethyl group, the tetrahydropyranyl group, the tert-butyl-dimethylsilyl group and the like.
-
- Reaction Scheme VI illustrates preparation of the conjugates provided herein of a peptidic substrate and the vinca alkaloid cytotoxic agent vinblastine wherein the attachment of vinblastine is at the C-terminus of the peptidic substrate. The use of the 1,3-diaminopropane linker is illustrative only; other linker units between the carbonyl of vinblastine and the C-terminus of the peptidic substrate are also envisioned (e.g., (CH2)u—T—(CH2)u). The acyl azide starting material is prepared from vinglasine by reaction with hydrazine (60-65° C., MeOH), followed by reaction with HCl/DMF/isoamyl nitrite. Furthermore, Reaction Scheme VI illustrates a synthesis of conjugates wherein the C4-hydroxy moiety is reacetylated following the addition of the linker unit. It is known that the desacetyl vinblastine conjugate also is efficacious and can be prepared by eliminating the steps shown in Reaction Scheme VI of protecting the primary amine of the linker and reacting the intermediate with acetic anhydride, followed by deprotection of the amine (see, e.g., International Patent Application Publication No. WO 98/10651). Conjugation of the peptidic substrate at other positions and functional groups of vinblastine can be readily accomplished by one of ordinary skill in the art and also is expected to provide conjugates that are substrates for cell surface proteases, or a soluble, shed or released form thereof.
- Reaction Scheme VII illustrates preparation of certain of the conjugates utilized in the compositions and methods provided herein wherein the peptidic substrates are combined with the vinca alkaloid cytotoxic agent vinblastine. Attachment of the N-terminus of the peptidic substrate to vinblastine is illustrated (S. P. Kandukuri et al. (1985)J. Med. Chem. 28:1079-1088).
- It also is understood that conjugates can be prepared wherein the N-terminus of the peptidic substrate utilized in the compositions and methods provided herein is combined with one therapeutic agent, such as a cytotoxic agent, such as vinblastine, while the C-terminus is simultaneously attached to another cytotoxic agent, which is the same or different cytotoxic agent, such as doxorubicin. Reaction Scheme VIII illustrates the synthesis of such a polycytotoxic agent conjugate. Such a polycytotoxic conjugate can offer advantages over a conjugate containing only one cytotoxic agent.
- With respect to the embodiment of a peptidic substrate combined with desacetylvinblastine, the following Reaction Schemes IX and X illustrate the synthesis of the conjugates provided herein.
- Reaction Scheme IX illustrates preparation of conjugates provided herein containing the peptidic substrates provided herein and the vinca alkaloid cytotoxic agent vinblastine wherein the attachment of the oxygen of the 4-desacetylvinblastine is at the C-terminus of the peptidic substrate. While other sequences of reactions can be useful in forming such conjugates, it is known that initial attachment of a single amino acid to the 4-oxygen and subsequent attachment of the remaining peptidic substrate sequence to that amino acid is an exemplarary method (see, International Patent Application Publication No. WO 99/28345). It also is known that 3,4-dihydro-3-hydroxy-4-oxo-1,2,3-benzotriazine (ODHBT) can be utilized in place of HOAt in the final coupling step.
-
-
- The conjugates and compositions provided herein are used for treating, preventing, or ameliorating one or more symptoms of any disease or disorder that can be treated by targeting a cell or tissue that expresses a cell surface protease, particularly, a serine protease, on its surface at higher levels compared to other cells, or soluble, shed or released forms thereof. These include, but are not limited to, hyperproliferative diseases, such as cancer, any disease associated with aberrant or excessive angiogenesis, autoimmune disorders, inflammatory diseases and any other disease for which an appropriate cell surface protease, including cell-associated and cell-localized proteases, can be identified.
- The pharmaceutical compositions provided herein contain therapeutically effective amounts of one or more of the conjugates provided herein that are useful in the prevention, treatment, or amelioration of one or more of the symptoms of diseases or disorders associated with undesired and/or uncontrolled angiogenesis or neovascularization. Such diseases or disorders include, but are not limited to, solid neoplasms, including lung, colon, esophageal, breast, ovarian and prostate cancers; vascular malformations and cardiovascular disorders, including, but not limited to, angiofibroma, angiolipoma, atherosclerosis, restenosis/reperfusion injury, arteriovenous malformations, hemangiomatosis and vascular adhesions, dyschondroplasia with vascular hematomas, hereditary hemorrhagic telangiectasia and Von Hipple Lindau syndrome; chronic inflammatory diseases and abherent wound repairs, including, but not limited to, diabetes mellitus, hemophiliac joints, inflammatory bowel disease, nonhealing fractures, rapidly progressing periodontitis, juvenile periodontitis, psoriasis, rheumatoid arthritis, venous stasis ulcers, granulations-burns, hypertrophic scars, liver cirrhosis, osteoradionecrosis, postoperative adhesions, pyogenic granuloma and systemic sclerosis; circulatory disorders, including, but not limited to, Raynaud's phenomenon; crest syndromes, including, but not limited to, calcinosis, esophageal, dyomotiloty, sclerodactyly and teangiectasis; dermatological disorders, including, but not limited to, systemic vasculitis, scleroderma, pyoderma gangrenosum, vasculopathy, venous, arterial ulcers, Sturge-Weber syndrome, Port-wine stains, blue rubber bleb nevus syndrome, Klippel-Trenaunay-Weber syndrome and Osler-Weber-Rendu syndrome; and ocular disorders, including, but not limited to, blindness caused by ocular neovascular disease, corneal graft neovascularization, macular degeneration in the eye, neovascular glaucoma, trachoma, diabetic retinopathy, myopic degeneration, retinopathy of prematurity, retrolental fibroplasia and corneal neovascularization.
- The compositions contain one or more conjugates provided herein. The conjugates can be formulated into suitable pharmaceutical preparations such as, for example, solutions, suspensions, tablets, dispersible tablets, pills, capsules, powders, sustained release formulations or elixirs, for oral administration or in sterile solutions or suspensions for parenteral administration, as well as transdermal patch preparation and dry powder inhalers. Typically the cojugates described above are formulated into pharmaceutical compositions using techniques and procedures well known in the art (see, e.g., Ansel (1985)Introduction to Pharmaceutical Dosage Forms, Fourth Edition, p. 126)). Effective concentrations can be empirically determined using animal models, in vitro models or test subjects.
- In the compositions, effective concentrations of one or more conjugates or pharmaceutically acceptable derivatives thereof is (are) mixed with a suitable pharmaceutical carrier or vehicle. The conjugates can be derivatized as the corresponding salts, esters, enol ethers or esters, acids, bases, solvates or hydrates prior to formulation, as described above. The concentrations of the conjugates in the compositions are effective for delivery of an amount, upon administration, that treats, prevents, or ameliorates one or more of the symptoms of diseases or disorders associated with undesired and/or uncontrolled angiogenesis or neovascularization. Such diseases or disorders include, but are not limited to, solid neoplasms; vascular malformations and cardiovascular disorders, including, but not limited to, angiofibroma, angiolipoma, atherosclerosis, restenosis/reperfusion injury, arteriovenous malformations, hemangiomatosis and vascular adhesions, dyschondroplasia with vascular hamartomas, hereditary hemorrhagic telangiectasia and Von Hipple Lindau syndrome; chronic inflammatory diseases and abherent wound repairs, including, but not limited to, diabetes mellitus, hemophiliac joints, inflammatory bowel disease, nonhealing fractures, rapidly progressing periodontitis, juvenile periodontitis, psoriasis, rheumatoid arthritis, venous stasis ulcers, granulations-burns, hypertrophic scars, liver cirrhosis, osteoradionecrosis, postoperative adhesions, pyogenic granuloma and systemic sclerosis; circulatory disorders, including, but not limited to, Raynaud's phenomenon; crest syndromes, including, but not limited to, calcinosis, esophageal, dyomotiloty, sclerodactyly and teangiectasis; dermatological disorders, including, but not limited to, systemic vasculitis, scleroderma, pyoderma gangrenosum, vasculopathy, venous, arterial ulcers, Sturge-Weber syndrome, Port-wine stains, blue rubber bleb nevus syndrome, Klippel-Trenaunay-Weber syndrome and Osler-Weber-Rendu syndrome; and ocular disorders, including, but not limited to, blindness caused by ocular neovascular disease, corneal graft neovascularization, macular degeneration in the eye, neovascular glaucoma, trachoma, diabetic retinopathy, myopic degeneration, retinopathy of prematurity, retrolental fibroplasia and corneal neovascularization.
- The conjugates herein can be formulated into pharmaceutical compositions suitable for topical, local, intravenous and systemic application. Effective concentrations of one or more of the conjugates are mixed with a suitable pharmaceutical carrier or vehicle. The concentrations or amounts of the conjugates that are effective requires delivery of an amount, upon administration, that ameliorates the symptoms or treats the disease. Typically, the compositions are formulated for single dosage administration. Therapeutically effective concentrations and amounts can be determined empirically by testing the conjugates in known in vitro and in vivo systems, such as those described here; dosages for humans or other animals can then be extrapolated therefrom.
- Upon mixing or addition of the conjugate(s) with the vehicle, the resulting mixture can be a solution, suspension, emulsion or other such composition. The form of the resulting mixture depends upon a number of factors, including the intended mode of administration and the solubility of the conjugate in the selected carrier or vehicle. The effective concentration is sufficient for ameliorating the symptoms of the disease, disorder or condition treated and can be empirically determined based upon in vitro and/or in vivo data, such as the data from the mouse xenograft model for tumors or rabbit ophthalmic model. If necessary, pharmaceutically acceptable salts or other derivatives of the conjugates can be prepared.
- Pharmaceutical carriers or vehicles suitable for administration of the conjugates provided herein include any such carriers known to those skilled in the art to be suitable for the particular mode of administration. In addition, the conjugates can be formulated as the sole pharmaceutically active ingredient in the composition or can be combined with other active ingredients.
- The conjugates can be administered by any appropriate route, for example, orally, parenterally, intravenously, intradermally, subcutaneously, or topically, in liquid, semi-liquid or solid form and are formulated in a manner suitable for each route of administration. Exemplary modes of administration depend upon the indication treated. Dermatological and ophthalmologic indications will typically be treated locally; whereas, tumors and vascular proliferative disorders, will typically be treated by systemic, intradermal or intramuscular, modes of administration.
- The conjugate is included in the pharmaceutically acceptable carrier in an amount sufficient to exert a therapeutically useful effect in the absence of undesirable side effects on the patient treated. It is understood that number and degree of side effects depends upon the condition for which the conjugates are administered. For example, certain toxic and undesirable side effects are tolerated when treating life-threatening illnesses, such as tumors, that would not be tolerated when treating disorders of lesser consequence.
- The concentration of conjugate in the composition will depend on absorption, inactivation and excretion rates thereof, the dosage schedule, and amount administered as well as other factors known to those of skill in the art.
- Typically a therapeutically effective dosage should produce a serum concentration of active ingredient of from about 0.1 ng/ml to about 50-100 μg/ml. The pharmaceutical compositions typically should provide a dosage of from about 0.01 mg to about 100-2000 mg of conjugate, depending upon the conjugate selected as adjusted for body surface area and/or weight. Typically, for intravenous or systemic treatment a daily dosage of about between 0.05 and 0.5 mg/kg should be sufficient. Local application for ophthalmic disorders should provide about 1 ng up to 100 μg, generally about 1 μg to about 10 μg, per single dosage administration. It is understood that the amount to administer is a function of the conjugate selected, the indication treated, and possibly the side effects that will be tolerated. Dosages can be empirically determined using recognized models for each disorder.
- Typically, the compositions are formulated for single dosage administration. To formulate a composition, the weight fraction of conjugate is dissolved, suspended, dispersed or otherwise mixed in a selected vehicle at an effective concentration such that the treated condition is relieved or ameliorated. Pharmaceutical carriers or vehicles suitable for administration of the conjugates provided herein include any such carriers known to those skilled in the art to be suitable for the particular mode of administration.
- In addition, the conjugates can be formulated as the sole ingredient in the composition or can be combined with other active ingredients. Liposomal suspensions, including tissue-targeted liposomes, particularly tumor-targeted liposomes, also can be suitable as pharmaceutically acceptable carriers. These can be prepared according to methods known to those skilled in the art. For example, liposome formulations can be prepared as described in U.S. Pat. No. 4,522,811. Briefly, liposomes such as multilamellar vesicles (MLV's) can be formed by drying down egg phosphatidyl choline and brain phosphatidyl serine (7:3 molar ratio) on the inside of a flask. A solution of a conjugate provided herein in phosphate buffered saline lacking divalent cations (PBS) is added and the flask shaken until the lipid film is dispersed. The resulting vesicles are washed to remove unencapsulated conjugate, pelleted by centrifugation, and then resuspended in PBS.
- The conjugate is included in the pharmaceutically acceptable carrier in an amount sufficient to exert a therapeutically useful effect in the absence of undesirable side effects on the patient treated. The therapeutically effective concentration can be determined empirically by testing the conjugates in in vitro and in vivo systems described herein (see, e.g., EXAMPLES 3 and 4) and then extrapolated therefrom for dosages for humans.
- The concentration of conjugate in the pharmaceutical composition will depend on absorption, inactivation and excretion rates of the conjugate, the physicochemical characteristics of the conjugate, the dosage schedule, and amount administered as well as other factors known to those of skill in the art. For example, the amount that is delivered is sufficient to ameliorate one or more of the symptoms of diseases or disorders associated with undesired and/or uncontrolled angiogenesis or neovascularization, as described herein.
- Typically a therapeutically effective dosage should produce a serum concentration of active ingredient of from about 0.1 ng/ml to about 50-100 μg/ml. The pharmaceutical compositions typically should provide a dosage of from about 0.001 mg to about 2000 mg of conjugate per kilo-gram of body weight per day. Pharmaceutical dosage unit forms are prepared to provide from about 1 mg to about 1000 mg and generally from about 10 to about 500 mg of the essential active ingredient or a combination of essential ingredients per dosage unit form.
- The conjugate can be administered at once, or can be divided into a number of smaller doses to be administered at intervals of time. It is understood that the precise dosage and duration of treatment is a function of the disease being treated and can be determined empirically using known testing protocols or by extrapolation from in vivo or in vitro test data. It is to be noted that concentrations and dosage values can also vary with the severity of the condition to be alleviated. It is to be further understood that for any particular subject, specific dosage regimens should be adjusted over time according to the individual need and the professional judgment of the person administering or supervising the administration of the compositions, and that the concentration ranges set forth herein are exemplary only and are not intended to limit the scope or practice of the claimed compositions.
- Exemplary pharmaceutically acceptable derivatives include acids, bases, enol ethers and esters, salts, esters, hydrates, solvates and conjugate forms. The derivative is selected such that its pharmacokinetic properties are superior to the corresponding neutral conjugate.
- Thus, effective concentrations or amounts of one or more of the conjugates described herein or pharmaceutically acceptable derivatives thereof are mixed with a suitable pharmaceutical carrier or vehicle for systemic, topical or local administration to form pharmaceutical compositions. Conjugates are included in an amount effective for ameliorating one or more symptoms of, or for treating or preventing diseases or disorders associated with undesired and/or uncontrolled angiogenesis or neovascularization, as described herein. The concentration of conjugate in the composition will depend on absorption, inactivation, excretion rates of the conjugate, the dosage schedule, amount administered, particular formulation as well as other factors known to those of skill in the art.
- The compositions are intended to be administered by a suitable route, including orally, parenterally, rectally, topically and locally. For oral administration, capsules and tablets are generally employed. The compositions are in liquid, semi-liquid or solid form and are formulated in a manner suitable for each route of administration. Exemplary modes of administration include parenteral and oral modes of administration.
- Solutions or suspensions used for parenteral, intradermal, subcutaneous, or topical application can include any of the following components: a sterile diluent, such as water for injection, saline solution, fixed oil, polyethylene glycol, glycerine, propylene glycol or other synthetic solvent; antimicrobial agents, such as benzyl alcohol and methyl parabens; antioxidants, such as ascorbic acid and sodium bisulfite; chelating agents, such as ethylenediaminetetraacetic acid (EDTA); buffers, such as acetates, citrates and phosphates; and agents for the adjustment of tonicity such as sodium chloride or dextrose. Parenteral preparations can be enclosed in ampules, disposable syringes or single or multiple dose vials made of glass, plastic or other suitable material.
- In instances in which the conjugates exhibit insufficient solubility, methods for solubilizing conjugates can be used. Such methods are known to those of skill in this art, and include, but are not limited to, using cosolvents, such as dimethylsulfoxide (DMSO), using surfactants, such as TWEEN®, or dissolution in aqueous sodium bicarbonate. Derivatives of the conjugates also can be used in formulating effective pharmaceutical compositions.
- Upon mixing or addition of the conjugate(s), the resulting mixture can be a solution, suspension, emulsion or the like. The form of the resulting mixture depends upon a number of factors, including the intended mode of administration and the solubility of the conjugate in the selected carrier or vehicle. The effective concentration is sufficient for ameliorating the symptoms of the disease, disorder or condition treated and can be empirically determined.
- The pharmaceutical compositions are provided for administration to humans and animals in unit dosage forms, such as tablets, capsules, pills, powders, granules, sterile parenteral solutions or suspensions, and oral solutions or suspensions, and oil-water emulsions containing suitable quantities of the conjugates or pharmaceutically acceptable derivatives thereof. The conjugates and derivatives thereof are typically formulated and administered in unit-dosage forms or multiple-dosage forms. Unit-dose forms as used herein refers to physically discrete units suitable for human and animal subjects and packaged individually as is known in the art. Each unit-dose contains a predetermined quantity of the conjugate sufficient to produce the desired therapeutic effect, in association with the required pharmaceutical carrier, vehicle or diluent. Examples of unit-dose forms include ampoules and syringes and individually packaged tablets or capsules. Unit-dose forms can be administered in fractions or multiples thereof. A multiple-dose form is a plurality of identical unit-dosage forms packaged in a single container to be administered in segregated unit-dose form. Examples of multiple-dose forms include vials, bottles of tablets or capsules or bottles of pints or gallons. Hence, multiple dose form is a multiple of unit-doses which are not segregated in packaging.
- The composition can contain along with the conjugate: a diluent such as lactose, sucrose, dicalcium phosphate, or carboxymethyl-cellulose; a lubricant, such as magnesium stearate, calcium stearate and talc; and a binder such as starch, natural gums, such as gum acacia-gelatin, glucose, molasses, polyvinylpyrrolidine, celluloses and derivatives thereof, povidone, crospovidones and other such binders known to those of skill in the art. Liquid pharmaceutically administrable compositions can, for example, be prepared by dissolving, dispersing, or otherwise mixing a conjugate as defined above and optional pharmaceutical adjuvants in a carrier, such as, for example, water, saline, aqueous dextrose, glycerol, glycols, ethanol, and the like, to thereby form a solution or suspension. If desired, the pharmaceutical composition to be administered can also contain minor amounts of nontoxic auxiliary substances such as wetting agents, emulsifying agents, or solubilizing agents, pH buffering agents and the like, for example, acetate, sodium citrate, cyclodextrine derivatives, sorbitan monolaurate, triethanolamine sodium acetate, triethanolamine oleate, and other such agents. Actual methods of preparing such dosage forms are known, or will be apparent, to those skilled in this art; for example, see Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, Pa., 15th Edition, 1975. The composition or formulation to be administered will, in any event, contain a quantity of the conjugate in an amount sufficient to alleviate the symptoms of the treated subject.
- Dosage forms or compositions containing active ingredient in the range of 0.005% to 100% with the balance made up from non-toxic carrier can be prepared. For oral administration, a pharmaceutically acceptable non-toxic composition is formed by the incorporation of any of the normally employed excipients, such as, for example pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, talcum, cellulose derivatives, sodium crosscarmellose, glucose, sucrose, magnesium carbonate or sodium saccharin. Such compositions include solutions, suspensions, tablets, capsules, powders and sustained release formulations, such as, but not limited to, implants and microencapsulated delivery systems, and biodegradable, biocompatible polymers, such as collagen, ethylene vinyl acetate, polyanhydrides, polyglycolic acid, polyorthoesters, polylactic acid and others. Methods for preparation of these compositions are known to those skilled in the art. The contemplated compositions can contain 0.001%-100% active ingredient, such as 0.1-85%, for example 75-95%.
- The conjugates or pharmaceutically acceptable derivatives can be prepared with carriers that protect the conjugate against rapid elimination from the body, such as time release formulations or coatings. The compositions can include other conjugates to obtain desired combinations of properties. The conjugates provided herein, or pharmaceutically acceptable derivatives thereof as described herein, also can be advantageously administered for therapeutic or prophylactic purposes together with another pharmacological agent known in the general art to be of value in treating one or more of the diseases or medical conditions referred to hereinabove, such as diseases or disorders associated with undesired and/or uncontrolled angiogenesis or neovascularization. It is to be understood that such combination therapy constitutes a further aspect of the compositions and methods of treatment provided herein.
- 1. Compositions for Oral Administration
- Oral pharmaceutical dosage forms are either solid, gel or liquid. The solid dosage forms are tablets, capsules, granules, and bulk powders. Types of oral tablets include compressed, chewable lozenges and tablets which can be enteric-coated, sugar-coated or film-coated. Capsules can be hard or soft gelatin capsules, while granules and powders can be provided in non-effervescent or effervescent form with the combination of other ingredients known to those skilled in the art.
- In certain embodiments, the formulations are solid dosage forms, such as, for example, capsules or tablets. The tablets, pills, capsules, troches and other dosage forms can contain, for example, any of the following ingredients, or compounds of a similar nature: a binder; a diluent; a disintegrating agent; a lubricant; a glidant; a sweetening agent; and a flavoring agent.
- Examples of binders include microcrystalline cellulose, gum tragacanth, glucose solution, acacia mucilage, gelatin solution, sucrose and starch paste. Lubricants include talc, starch, magnesium or calcium stearate, lycopodium and stearic acid. Diluents include, for example, lactose, sucrose, starch, kaolin, salt, mannitol and dicalcium phosphate. Glidants include, but are not limited to, colloidal silicon dioxide. Disintegrating agents include crosscarmellose sodium, sodium starch glycolate, alginic acid, corn starch, potato starch, bentonite, methylcellulose, agar and carboxymethylcellulose. Coloring agents include, for example, any of the approved certified water soluble FD and C dyes, mixtures thereof; and water insoluble FD and C dyes suspended on alumina hydrate. Sweetening agents include sucrose, lactose, mannitol and artificial sweetening agents such as saccharin, and any number of spray dried flavors. Flavoring agents include natural flavors extracted from plants such as fruits and synthetic blends of conjugates which produce a pleasant sensation, such as, but not limited to peppermint and methyl salicylate. Wetting agents include propylene glycol monostearate, sorbitan monooleate, diethylene glycol monolaurate and polyoxyethylene laural ether. Emetic-coatings include fatty acids, fats, waxes, shellac, ammoniated shellac and cellulose acetate phthalates. Film coatings include hydroxyethylcellulose, sodium carboxymethylcellulose, polyethylene glycol 4000 and cellulose acetate phthalate.
- If oral administration is desired, the conjugate could be provided in a composition that protects it from the acidic environment of the stomach. For example, the composition can be formulated in an enteric coating that maintains its integrity in the stomach and releases the conjugate in the intestine. The composition also can be formulated in combination with an antacid or other such ingredient.
- When the dosage unit form is a capsule, it can contain, in addition to material of the above type, a liquid carrier such as a fatty oil. In addition, dosage unit forms can contain various other materials which modify the physical form of the dosage unit, for example, coatings of sugar and other enteric agents. The conjugates also can be administered as a component of an elixir, suspension, syrup, wafer, sprinkle, chewing gum or the like. A syrup can contain, in addition to the conjugates, sucrose as a sweetening agent and certain preservatives, dyes and colorings and flavors.
- The conjugates also can be mixed with other active materials which do not impair the desired action, or with materials that supplement the desired action, such as antacids, H2 blockers, and diuretics. Higher concentrations, up to about 98% by weight of the conjugate can be included.
- Pharmaceutically acceptable carriers included in tablets are binders, lubricants, diluents, disintegrating agents, coloring agents, flavoring agents, and wetting agents. Enteric-coated tablets, because of the enteric-coating, resist the action of stomach acid and dissolve or disintegrate in the neutral or alkaline intestines. Sugar-coated tablets are compressed tablets to which different layers of pharmaceutically acceptable substances are applied. Film-coated tablets are compressed tablets which have been coated with a polymer or other suitable coating. Multiple compressed tablets are compressed tablets made by more than one compression cycle utilizing the pharmaceutically acceptable substances previously mentioned. Coloring agents also can be used in the above dosage forms. Flavoring and sweetening agents are used in compressed tablets, sugar-coated, multiple compressed and chewable tablets. Flavoring and sweetening agents are especially useful in the formation of chewable tablets and lozenges.
- Liquid oral dosage forms include aqueous solutions, emulsions, suspensions, solutions and/or suspensions reconstituted from non-effervescent granules and effervescent preparations reconstituted from effervescent granules. Aqueous solutions include, for example, elixirs and syrups. Emulsions are either oil-in-water or water-in-oil.
- Elixirs are clear, sweetened, hydroalcoholic preparations. Pharmaceutically acceptable carriers used in elixirs include solvents. Syrups are concentrated aqueous solutions of a sugar, for example, sucrose, and can contain a preservative. An emulsion is a two-phase system in which one liquid is dispersed in the form of small globules throughout another liquid. Pharmaceutically acceptable carriers used in emulsions are non-aqueous liquids, emulsifying agents and preservatives. Suspensions use pharmaceutically acceptable suspending agents and preservatives. Pharmaceutically acceptable substances used in non-effervescent granules, to be reconstituted into a liquid oral dosage form, include diluents, sweeteners and wetting agents. Pharmaceutically acceptable substances used in effervescent granules, to be reconstituted into a liquid oral dosage form, include organic acids and a source of carbon dioxide. Coloring and flavoring agents are used in all of the above dosage forms.
- Solvents include glycerin, sorbitol, ethyl alcohol and syrup. Examples of preservatives include glycerin, methyl and propylparaben, benzoic add, sodium benzoate and alcohol. Examples of non-aqueous liquids utilized in emulsions include mineral oil and cottonseed oil. Examples of emulsifying agents include gelatin, acacia, tragacanth, bentonite, and surfactants such as polyoxyethylene sorbitan monooleate. Suspending agents include sodium carboxymethylcellulose, pectin, tragacanth, Veegum and acacia. Diluents include lactose and sucrose. Sweetening agents include sucrose, syrups, glycerin and artificial sweetening agents such as saccharin. Wetting agents include propylene glycol monostearate, sorbitan monooleate, diethylene glycol monolaurate and polyoxyethylene lauryl ether. Organic adds include citric and tartaric acid. Sources of carbon dioxide include sodium bicarbonate and sodium carbonate. Coloring agents include any of the approved certified water soluble FD and C dyes, and mixtures thereof. Flavoring agents include natural flavors extracted from plants such fruits, and synthetic blends of conjugates which produce a pleasant taste sensation.
- For a solid dosage form, the solution or suspension, in for example propylene carbonate, vegetable oils or triglycerides, for example the formulation can be encapsulated in a gelatin capsule. Such solutions, and the preparation and encapsulation thereof, are disclosed in U.S. Pat. Nos. 4,328,245; 4,409,239; and 4,410,545. For a liquid dosage form, the solution, e.g., for example, in a polyethylene glycol, can be diluted with a sufficient quantity of a pharmaceutically acceptable liquid carrier, e.g., water, to be easily measured for administration.
- Alternatively, liquid or semi-solid oral formulations can be prepared by dissolving or dispersing the conjugate or derivative thereof in vegetable oils, glycols, triglycerides, propylene glycol esters (e.g., propylene carbonate) and other such carriers, and encapsulating these solutions or suspensions in hard or soft gelatin capsule shells. Other useful formulations include those set forth in U.S. Pat. Nos. Re 28,819 and 4,358,603. Briefly, such formulations include, but are not limited to, those containing a conjugate provided herein, a dialkylated mono- or poly-alkylene glycol, including, but not limited to, 1,2-dimethoxymethane, diglyme, triglyme, tetraglyme, polyethylene glycol-350-dimethyl ether, polyethylene glycol-550-dimethyl ether, polyethylene glycol-750-dimethyl ether wherein 350, 550 and 750 refer to the approximate average molecular weight of the polyethylene glycol, and one or more anitoxidants, such as butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), propyl gallate, vitamin E, hydroquinone, hydroxycoumarins, ethanolamine, lecithin, cephalin, ascorbic acid, malic acid, sorbitol, phosphoric acid, thiodipropionic acid and its esters, and dithiocarbamates.
- Other formulations include, but are not limited to, aqueous alcoholic solutions including a pharmaceutically acceptable acetal. Alcohols used in these formulations are any pharmaceutically acceptable water-miscible solvents having one or more hydroxyl groups, including, but not limited to, propylene glycol and ethanol. Acetals include, but are not limited to, di(lower alkyl) acetals of lower alkyl aldehydes such as acetaldehyde diethyl acetal.
- In all embodiments, tablets and capsules formulations can be coated as known by those of skill in the art in order to modify or sustain dissolution of the conjugate. Thus, for example, they can be coated with a conventional enterically digestible coating, such as phenylsalicylate, waxes and cellulose acetate phthalate.
- 2. Injectables, Solutions and Emulsions
- Parenteral administration, generally characterized by injection, either subcutaneously, intramuscularly or intravenously also is contemplated herein. Injectables can be prepared in conventional forms, either as liquid solutions or suspensions, solid forms suitable for solution or suspension in liquid prior to injection, or as emulsions. Suitable excipients are, for example, water, saline, dextrose, glycerol or ethanol. In addition, if desired, the pharmaceutical compositions to be administered can also contain minor amounts of non-toxic auxiliary substances such as wetting or emulsifying agents, pH buffering agents, stabilizers, solubility enhancers, and other such agents, such as for example, sodium acetate, sorbitan monolaurate, triethanolamine oleate and cyclodextrins. Implantation of a slow-release or sustained-release system, such that a constant level of dosage is maintained (see, e.g., U.S. Pat. No. 3,710,795) also is contemplated herein. Briefly, a conjugate provided herein is dispersed in a solid inner matrix, e.g., polymethylmethacrylate, polybutylmethacrylate, plasticized or unplasticized polyvinylchloride, plasticized nylon, plasticized polyethyleneterephthalate, natural rubber, polyisoprene, polyisobutylene, polybutadiene, polyethylene, ethylene-vinylacetate copolymers, silicone rubbers, polydimethylsiloxanes, silicone carbonate copolymers, hydrophilic polymers such as hydrogels of esters of acrylic and methacrylic acid, collagen, cross-linked polyvinylalcohol and cross-linked partially hydrolyzed polyvinyl acetate, that is surrounded by an outer polymeric membrane, e.g., polyethylene, polypropylene, ethylene/propylene copolymers, ethylene/ethyl acrylate copolymers, ethylene/vinylacetate copolymers, silicone rubbers, polydimethyl siloxanes, neoprene rubber, chlorinated polyethylene, polyvinylchloride, vinylchloride copolymers with vinyl acetate, vinylidene chloride, ethylene and propylene, ionomer polyethylene terephthalate, butyl rubber epichlorohydrin rubbers, ethylene/vinyl alcohol copolymer, ethylene/vinyl acetate/vinyl alcohol terpolymer, and ethylene/vinyloxyethanol copolymer, that is insoluble in body fluids. The conjugate diffuses through the outer polymeric membrane in a release rate controlling step. The percentage of conjugate contained in such parenteral compositions is highly dependent on the specific nature thereof, as well as the activity of the conjugate and the needs of the subject.
- Parenteral administration of the compositions includes intravenous, subcutaneous and intramuscular administrations. Preparations for parenteral administration include sterile solutions ready for injection, sterile dry soluble products, such as lyophilized powders, ready to be combined with a solvent just prior to use, including hypodermic tablets, sterile suspensions ready for injection, sterile dry insoluble products ready to be combined with a vehicle just prior to use and sterile emulsions. The solutions can be either aqueous or nonaqueous.
- If administered intravenously, suitable carriers include physiological saline or phosphate buffered saline (PBS), and solutions containing thickening and solubilizing agents, such as glucose, polyethylene glycol, and polypropylene glycol and mixtures thereof.
- Pharmaceutically acceptable carriers used in parenteral preparations include aqueous vehicles, nonaqueous vehicles, antimicrobial agents, isotonic agents, buffers, antioxidants, local anesthetics, suspending and dispersing agents, emulsifying agents, sequestering or chelating agents and other pharmaceutically acceptable substances.
- Examples of aqueous vehicles include Sodium Chloride Injection, Ringers Injection, Isotonic Dextrose Injection, Sterile Water Injection, Dextrose and Lactated Ringers Injection. Nonaqueous parenteral vehicles include fixed oils of vegetable origin, cottonseed oil, corn oil, sesame oil and peanut oil. Antimicrobial agents in bacteriostatic or fungistatic concentrations must be added to parenteral preparations packaged in multiple-dose containers which include phenols or cresols, mercurials, benzyl alcohol, chlorobutanol, methyl and propyl p-hydroxybenzoic acid esters, thimerosal, benzalkonium chloride and benzethonium chloride. Isotonic agents include sodium chloride and dextrose. Buffers include phosphate and citrate. Antioxidants include sodium bisulfate. Local anesthetics include procaine hydrochloride. Suspending and dispersing agents include sodium carboxymethylcelluose, hydroxypropyl methylcellulose and polyvinylpyrrolidone. Emulsifying agents include Polysorbate 80 (TWEEN® 80). A sequestering or chelating agent of metal ions include EDTA. Pharmaceutical carriers also include ethyl alcohol, polyethylene glycol and propylene glycol for water miscible vehicles and sodium hydroxide, hydrochloric acid, citric acid or lactic acid for pH adjustment.
- The concentration of the conjugate is adjusted so that an injection provides an effective amount to produce the desired pharmacological effect. The exact dose depends on the age, weight and condition of the patient or animal as is known in the art.
- The unit-dose parenteral preparations are packaged in an ampule, a vial or a syringe with a needle. All preparations for parenteral administration must be sterile, as is known and practiced in the art.
- Illustratively, intravenous or intraarterial infusion of a sterile aqueous solution containing a conjugate is an effective mode of administration. Another embodiment is a sterile aqueous or oily solution or suspension containing a conjugate injected as necessary to produce the desired pharmacological effect.
- Injectables are designed for local and systemic administration. Typically a therapeutically effective dosage is formulated to contain a concentration of at least about 0.1% w/w up to about 90% w/w or more, genrally more than 1% w/w of the conjugate to the treated tissue(s). The conjugate can be administered at once, or can be divided into a number of smaller doses to be administered at intervals of time. It is understood that the precise dosage and duration of treatment is a function of the tissue being treated and can be determined empirically using known testing protocols or by extrapolation from in vivo or in vitro test data. It is to be noted that concentrations and dosage values can also vary with the age of the individual treated. It is to be further understood that for any particular subject, specific dosage regimens should be adjusted over time according to the individual need and the professional judgment of the person administering or supervising the administration of the formulations, and that the concentration ranges set forth herein are exemplary only and are not intended to limit the scope or practice of the claimed formulations.
- The conjugate can be suspended in micronized or other suitable form or can be derivatized to produce a more soluble product. The form of the resulting mixture depends upon a number of factors, including the intended mode of administration and the solubility of the conjugate in the selected carrier or vehicle. The effective concentration is sufficient for ameliorating the symptoms of the condition and can be empirically determined.
- 3. Lyophilized Powders
- Of interest herein are also lyophilized powders, which can be reconstituted for administration as solutions, emulsions and other mixtures. They also can be reconstituted and formulated as solids or gels.
- The sterile, lyophilized powder is prepared by dissolving a conjugate provided herein, or a pharmaceutically acceptable derivative thereof, in a suitable solvent. The solvent can contain an excipient which improves the stability or other pharmacological component of the powder or reconstituted solution, prepared from the powder. Excipients that can be used include, but are not limited to, dextrose, sorbital, fructose, corn syrup, xylitol, glycerin, glucose, sucrose or other suitable agent. The solvent can also contain a buffer, such as citrate, sodium or potassium phosphate or other such buffer known to those of skill in the art at, typically, about neutral pH. Subsequent sterile filtration of the solution followed by lyophilization under standard conditions known to those of skill in the art provides the desired formulation. Generally, the resulting solution will be apportioned into vials for lyophilization. Each vial will contain a single dosage (such as 10-1000 mg, for example 100-500 mg) or multiple dosages of the conjugate. The lyophilized powder can be stored under appropriate conditions, such as at about 4° C. to room temperature.
- Reconstitution of this lyophilized powder with water for injection provides a formulation for use in parenteral administration. For reconstitution, generally about 1-50 mg, such 5-35 mg or about 9-30 mg of lyophilized powder, is added per mL of sterile water or other suitable carrier. The precise amount depends upon the selected conjugate, intended subject, and other empircally determinable parameters. Hence the amount can be empirically determined.
- 4. Topical Administration
- Topical mixtures are prepared as described for the local and systemic administration. The resulting mixture can be a solution, suspension, emulsions or the like and are formulated as creams, gels, ointments, emulsions, solutions, elixirs, lotions, suspensions, tinctures, pastes, foams, aerosols, irrigations, sprays, suppositories, bandages, dermal patches or any other formulations suitable for topical administration.
- The conjugates or pharmaceutically acceptable derivatives thereof can be formulated as aerosols for topical application, such as by inhalation (see, e.g., U.S. Pat. Nos. 4,044,126, 4,414,209, and 4,364,923, which describe aerosols for delivery of a steroid useful for treatment of inflammatory diseases, particularly asthma). These formulations for administration to the respiratory tract can be in the form of an aerosol or solution for a nebulizer, or as a microfine powder for insufflation, alone or in combination with an inert carrier such as lactose. In such a case, the particles of the formulation will typically have diameters of less than 50 microns, generally less than 10 microns.
- The conjugates can be formulated for local or topical application, such as for topical application to the skin and mucous membranes, such as in the eye, in the form of gels, creams, and lotions and for application to the eye or for intracisternal or intraspinal application. Topical administration is contemplated for transdermal delivery and also for administration to the eyes or mucosa, or for inhalation therapies. Nasal solutions of the conjugate alone or in combination with other pharmaceutically acceptable excipients also can be administered.
- These solutions, particularly those intended for ophthalmic use, can be formulated as 0.01%-10% isotonic solutions, pH about 5-7, with appropriate salts.
- 5. Compositions for other Routes of Administration
- Other routes of administration, such as topical application, transdermal patches, and rectal administration are also contemplated herein.
- For example, pharmaceutical dosage forms for rectal administration are rectal suppositories, capsules and tablets for systemic effect. Rectal suppositories are used herein mean solid bodies for insertion into the rectum which melt or soften at body temperature releasing one or more conjugates. Pharmaceutically acceptable substances utilized in rectal suppositories are bases or vehicles and agents to raise the melting point. Examples of bases include cocoa butter (theobroma oil), glycerin-gelatin, carbowax (polyoxyethylene glycol) and appropriate mixtures of mono-, di- and triglycerides of fatty acids. Combinations of the various bases can be used. Agents to raise the melting point of suppositories include spermaceti and wax. Rectal suppositories can be prepared either by the compressed method or by molding. The typical weight of a rectal suppository is about 2 to 3 gm.
- Tablets and capsules for rectal administration are manufactured using the same pharmaceutically acceptable substance and by the same methods as for formulations for oral administration.
- 6. Articles of Manufacture
- The conjugates or pharmaceutically acceptable derivatives can be packaged as articles of manufacture containing packaging material, a conjugate or pharmaceutically acceptable derivative thereof provided herein, which is used for treatment, prevention or amelioration of one or more symptoms associated with proliferative diseases or disorders, and a label that indicates that the conjugate or pharmaceutically acceptable derivative thereof is used for treatment, prevention or amelioration of one or more symptoms associated with proliferative diseases or disorders.
- The articles of manufacture provided herein contain packaging materials. Packaging materials for use in packaging pharmaceutical products are well known to those of skill in the art. See, e.g., U.S. Pat. Nos. 5,323,907, 5,052,558 and 5,033,252. Examples of pharmaceutical packaging materials include, but are not limited to, blister packs, bottles, tubes, inhalers, pumps, bags, vials, containers, syringes, bottles, and any packaging material suitable for a selected formulation and intended mode of administration and treatment. A wide array of formulations of the conjugates and compositions provided herein are contemplated as are a variety of treatments for any disorder in which a cell surface protease, or a soluble, shed or secreted form thereof, is implicated.
- Standard physiological, pharmacological and biochemical procedures are available for testing the conjugates to identify those that possess therapeutic activity upon action of a cell surface protease or a soluble, shed, or released form thereof. In vitro and in vivo assays that can be used to evaluate therapeutic activity, such as cytotoxicity, of the conjugates will depend upon the therapeutic agent being tested.
- Exemplary assays are discussed briefly below with reference to cytotoxic conjugates (see, also, Examples). It is understood that the particular activity assayed will depend upon the conjugated therapeutic agent.
- 1. In Vitro Assays
- The therapeutic activity, such as cytotoxicity, of the conjugates provided herein can assessed by any assays normally used for assessing the therapeutic activity, such as cytotoxicity, of the unconjugated therapeutic agent. Numerous such assays are known, for example, assays can employ cells that express the targeted cell surface protease and the therapeutic activity of the therapeutic agent is assessed. For example, cytoxicity can be assessed by measuring cell viability or by measuring cell proliferation, such as by incorporation of a labeled nucleotide or other such label. Generally the activity is compared with cells that do not express the targeted protease.
- For example, the cells will be any that express a targeted MTSP or endotheliase. Such cells can be obtained by choosing cells known to express the cell surface protease, such as by determining tissue expression profiles, as discussed above, or by screening a variety of cell lines with an antibody for a targeted protease, or for the protease activity in the presence of a labeled, such as a chromogenic, substrate for the protease in the presence and absence of a known inhibitor of the targeted protease.
- Alternatively, nucleic acid encoding the protease can be introduced in a cell line that does not express the protease, and expressed therein to produce a cell line that expresses the protease of interest. The resulting recombinant cells can be used in cytotoxicity assays.
- 2. In Vivo Assays
- Numerous animal models for assessing therapeutic activity are known. Any suitable in vivo model can be used. Exemplary are the mouse xenograft model and chicken embryo models.
- Chicken Embryo Model
- The CAM model (chick embryo chorioallantoic membrane model; Ossowski (1988)J. Cell Biol. 107:2437-2445), provides another method for evaluating the inhibitory activity of a test compound. In the CAM model, tumor cells invade through the chorioallantoic membrane containing CAM (with tumor cells in the presence of several serine protease inhibitors results in less or no invasion of the tumor cells through the membrane). Thus, the CAM assay is performed with CAM and tumor cells in the presence and absence of various concentrations of test compound. The invasiveness of tumor cells is measured under such conditions to provide an indication of the compound's inhibitory activity. A compound having inhibitory activity correlates with less tumor invasion.
- Thus, the CAM assay is performed with CAM and tumor cells in the presence and absence of various concentrations of a test compound. A compound having activity correlates with a change in tumor invasion and/or tumor growth.
- For example, the ability of a cell surface protease to liberate a therapeutic agent, such as a cytotoxic agent, or the activity of a conjugate agent can be assessed using this model. If the therapeutic agent is released from the compound and it is an inhibitory agent there will be less tumor invasion or a decrease in size of the tumor. If the therapeutic agent is inactive in the conjugate, there will be no effect on tumor invasion.
- The CAM model also is used in a standard assay of angiogenesis (i.e., effect on formation of new blood vessels (Brooks et al. (1991)Methods in Molecular Biology 129:257-269). According to this model, a filter disc containing an angiogenesis inducer, such as basic fibroblast growth factor (bFGF) is placed onto the CAM. Diffusion of the cytokine into the CAM induces local angiogenesis, which can be measured in several ways such as by counting the number of blood vessel branch points within the CAM directly below the filter disc. The ability of identified compounds to inhibit cytokine-induced angiogenesis can be tested using this model. A test compound can either be added to the filter disc that contains the angiogenesis inducer, be placed directly on the membrane or be administered systemically. The extent of new blood vessel formation in the presence and/or absence of test compound can be compared using this model. The formation of fewer new blood vessels in the presence of a test compound would be indicative of anti-angiogenesis activity.
- This can be adapted for use with the conjugates herein to 1) assess the activity of a therapeutic agent in the conjugate; and 2) to assess the ability of a particular cell surface protease to liberate a therapeutic agent from a conjugate.
- Mouse Xenograft Model
- In vivo activity can be a assessed using recognized animal models, such as the well-known mouse xenograft model for anti-tumor activity (see, e.g., Beitz et al. (1992)Cancer Research 52:227-230; Houghton et al. (1982) Cancer Res. 42:535-539; Bogden et al. (1981) Cancer (Philadelphia) 48:10-20; Hoogenhout et al. (1983) Int. J. Radiat. Oncol., Biol. Phys. 9:871-879; Stastny et al. (1993) Cancer Res. 53:5740-5744). The in vivo mouse solid tumor xenograft model is used in assays for that test an agent's ability to inhibit tumor cell proliferation and/or spontaneous metastasis. For example, a conjugate is evaluated for anti-tumor activity against any tumor subtype that expresses the targeted cell surface protease, e.g., an ovarian tumor, in a mouse tumor xenograft model. Nude mice are given one or more, such as four intravenous injections of the conjugate. Dosing material is prepared by mixing the test material with appropriate volumes of, for example, PBS/0.1% BSA to achieve the desired doses. Mice IV injections (250-300 ul) into the tail vein for the duration of the experiment, such as, for example, days 5, 12, 19 and 26, with
day 1 designated as the day that the tumor cells are injected into the mice. Doses are either fixed or normalized for differences in body weight. Tumor volume is measured twice weekly for a selected period. - Female Balb/c nu/nu athymic mice (Roger Williams Hospital Animal Facility, Providence, R.I.), 8-12 weeks old are suitable mice. They should be maintained in an aseptic environment and selected such that body weights range from about 25-30 grams the day prior to dosing. Animals are maintained in a quarantined room and handled under aseptic conditions. Food and water are supplied ad libitum. Appropriate tumor cells can be obtained, for example, from the American Type Culture Collection (Rockville, Md.) and grown in modified Eagle's medium supplemented with 10% fetal calf serum. A selected number of days, such as five days prior to injection of the test material, mice receive a subcutaneous injection of tumor cells in the right rear flank.
- Calipers are used to measure the dimensions of each tumor. Measurements (mm) of maximum and minimum width are performed prior to injection of the test material and at selected, such as bi-weekly, intervals for the duration of the experiment. Tumor volumes (mm3) can be computed, for example, using the formula:
- Volume=[(width)2(length)]/2.
- Also provided are methods for identifying proteases to target conjugates for treatment of diseases. The methods involve identifying cell-surface protease-associated disease by identifying a cell involved in the disease process or a cell in the vicinity of the cell involved in the disease process. For example, if disease involves a particular tumor, a protease present on the particular tumor or on cells that a located in the vicinity thereof is identified. A cell surface protease on the cell for targeting and substrates therefor are then identified. Conjugates that target such proteases as provided herein can then be prepared.
- The following examples are included for illustrative purposes only and are not intended to limit the scope of the invention.
- Step A: Synthesis of Peptides on Wang Resin
- Peptides were prepared automatically using an ABI 431A peptide synthesizer from Perseptive Biosystems on preloaded Wang resin (0.25 mmol). The ABI 431A uses HOBT, HBTU, DIEA activation. The synthesis of N-acetyl (or other amide) capped peptides involved the use of AcOH (or other respective carboxylic acid) during the final coupling step on the ABI 431A. Other N-terminal caps where attached manually by using the following reagents: For carbamates and sulfonamides the peptides were capped with ROCOCl or RSO2Cl and DIEA (4 equivalents each, 1 hr) in DMF (3 mL).
- Step B: Cleavage of Peptides from Wang Resin
- The cleavage of peptides from Wang resin involved shaking the resin with 2 mL TFA/H2O (95:5) for 45 min. The resin was removed by filtration and the filtrate was allowed to stand for an additional hour. The solution was concentrated to a residue. The crude peptide was analyzed by analytical HPLC (system A). Typical purity of the crude peptide ranged from 80% to 95%. The peptides were purified by preparative HPLC (system B) using an appropriate gradient (typically 10-30%). Pure fractions were then lyophilized to provide the desired peptide as a white solid. Typical yields were 20-50% and a purity of 96-99%.
- Analytical HPLC Conditions (System A)
- Column: Chromolith RP-18e 4.6 mm×100 mm from EM science
- Gradient: 5-50% B in A over 6 min
- Flow Rate: 4 mL/min
- Solvent A: 0.1% TFA in water
- Solvent B: 0.1% TFA in acetonitrile
- Wavelength: 210 nm, 280 nm
- Preparative HPLC Conditions (System B)
- Column: Ultro 120 5 C18Q 150×20 mm from Peeke Scientific
- Gradient: 0-20%, or 10-30% or 20-40% B in A over 40 min
- Flow Rate: 18 mL/min
- Solvent A: 0.1% TFA in water
- Solvent B: acetonitrile
- Wavelength: 214 nm
- Step C: Coupling of Peptide acids to Doxorubicin
- To a mixture of peptide acid (0.052 mmol, 1.2 equivalents), doxorubicin hydrochloride (0.043 mmol, 25 mg), and HATU (0.0604 mmol, 22.9 mg, 1.4 equivalents) was added DMF (1 mL) then 2,6-lutidine (0.17 mmol, 20 μL, 4 eqiuvalents). The mixture was mixed until a homogeneous solution was obtained. After 4 to 24 hours (monitor by HPLC system A) the reaction was diluted with water (9 mL) and directly purified by preparative HPLC (system D). Pure fractions were then lyophilized to provide the desired peptide doxorubicin conjugate as a fluffy red solid. The quality of the final conjugate was verified by analytical HPLC (system C) and mass spectroscopy. Typical yields were 10-30% with a purity of 95-99%. (Note: when the peptide acid contained a histidine residue DIEA was substituted as the base and the reaction time was shortened to 1 hour).
- Deprotection of fluorenylmethylesters of peptide doxorubicin conjugates: In cases where free carboxylic acid is present in the conjugate a fluorenyl methyl ester was used to protect a carboxylic acid during coupling of the C-terminus of the peptide acid to doxorubicin, the flourenylmethyl group was subsequently removed with 10% morpholine in DMF for 1 hour.
- Analytical HPLC Conditions (System C)
- Column: Chromolith RP-18e 4.6 mm×100 mm from EM science
- Gradient: 5-50% B in A over 6 min
- Flow Rate: 4 mL/min
- Solvent A: 0.1% TFA in water
- Solvent B: 0.1% TFA in acetonitrile
- Wavelength: 210 nm, 280 nm
- Examples of retention times (min)
Doxorubicin 4.05 Ac-Gly-Ser-Gly-Arg-Ser-nLeu-Dox 4.34 MeOCO-Thr-Gly-Arg-Ser-nLeu-Dox 4.39 PhSO2-Thr-Gly-Arg-Ser-nLeu-Dox 4.83 N,N-dimethylglycine-Thr-Gly-Arg-Ser-nLeu-Dox 4.27 Ac-Thr-Gly-Arg-Ser-nLeu-Dox 4.32 - Preparative HPLC Conditions (System D)
- Column: Ultro 120 5 C18Q 150×20 mm from Peeke Scientific
- Gradient: 10-30% B in A over 40 min
- Flow Rate: 18 mL/min
- Solvent A: 0.1% acetic acid in water
- Solvent B: acetonitrile
- Wavelength: 214 nm
- Step A: Manual Synthesis of Ac-Gly-Ser(tBu)-Gly-Arg(Pbf)-Ser(tBu)-nLeu-Wang Resin
- In a 250 mL fritted peptide synthesis vessel equipped with nitrogen agitation and vacumm assisted drainage, Fmoc-nL-Wang resin (nova-biochem, 3.3 grams, 0.9 mmol/g, 3 mmol) was pre-swelled for 30 min using DMF. The peptide was then elongated by repeating the 4 step procedure below a total of five times with the following Fmoc aminoacids: Fmoc-Ser(tBu)-OH, Fmoc-Arg(Pbf)-OH, Fmoc-Gly-OH, Fmoc-Ser(tBu)-OH, Fmoc-Gly-OH.
- Iterative Coupling Procedure
- 1. the resin was mixed with 20% piperdine in DMF (100 mL) for 5 min then drained (
repeat 3 times). - 2. the resin was agitated with DMF (100 mL) for 30 sec then drained (
repeat 3 times). - 3. to a mixture of Fmoc-aminoacid (12 mmol), HOBT (12 mmol, 4 equivalents, 1.622 g), TBTU (11.7 mmol, 3.9 equivalents, 3.757 g), DMF (10 mL) and NMP (90 mL) was added DIEA (12 mmol, 4 equivalents, 2.10 mL). After stirring for 5 min to allow pre-activation, the solution was added to the synthesis vessel. The reaction was checked for completion by ninhydrin test and then drained. (If the ninhydrid test was blue, a double coupling (repeat step 3) was performed.
- 4. the resin was agitated with DMF (100 mL) for 30 sec then drained (
repeat 3 times). - The elongated resin (Fmoc-Gly-Ser(tBu)-Gly-Arg(Pbf)-Ser(tBu)-nLeu-Wang resin) was treated to
steps - Step B: Preparation of Ac-Gly-Ser-Gly-Arg-Ser-nLeu-OH
- To the above synthesis vessel containing Ac-Gly-Ser(tBu)-Gly-Arg(Pbf)-Ser(tBu)-nLeu-Wang resin (3 mmol) was added TFA/H2O (95:5, 50 mL). After gently agitation for 45 min the cleavage solution was collected and the filtrate was allowed to stand for an additional 90 min. The solution was concentrated to a residue. The crude peptide was analyzed by analytical HPLC (system A, RT=1.73, purity=90%). The residue was dissolved in water (50 mL) and hexanes (10 mL) and mixed. The hexanes layer was removed and the aqueous layer bubbled with nitrogen to evaporate any remaining hexanes. The crude peptide was purified by preparative HPLC (system E). Pure fractions were then lyophilized to provide Ac-Gly-Ser-Gly-Arg-Ser-nLeu-OH (1.04 g, 1.68 mmol, 56%) as a white solid. The purity was evaluated by analytical HPLC (system A, RT=1.73 min, 97% purity) and the constitution by mass spectrospcopy (ion observed at 617.9).
- Preparative HPLC Conditions (System E)
- Column: Waters Delta-Pak radial compression column, 15 um, 100A
- Gradient: 5-15% B in A over 40 min
- Flow Rate: 80 mL/min
- Solvent A: 0.1% acetic acid in water
- Solvent B: acetonitrile
- Wavelength: 214 nm
- Step C: Preparation of Ac-Gly-Ser-Gly-Arg-Ser-nLeu-Dox
- To a mixture of Ac-Gly-Ser-Gly-Arg-Ser-nLeu-OH (1.68 mmol, 1.04 g, 1.1 equivalents), doxorubicin hydrochloride (1.53 mmol, 887.8 mg), and HATU (1.76 mmol, 669.6 mg, 1.15 equivalents) was added DMF (40 mL) then 2,6-lutidine (6.12 mmol, 709 μL, 4 eqiuvalents). The solution was stirred for 18 hours. The reaction was diluted with water (100 mL), acidified with acetic acid (400 μL) and purified immediately in three batches by preparative HPLC (system E). Each red colored fraction was analyzed by analytical HPLC (system F). Fractions of greater than 95% purity were then combined. The acetonitrile was removed under vacuum and the remaining solution was lyophilized to provide Ac-Gly-Ser-Gly-Arg-Ser-nLeu-Dox (0.682 mmol, 780 mg, 45%) as a fluffy red solid. The purity was evaluated by analytical HPLC (system F, RT=3.51 min, 95% purity) and the constitution by mass spectrospcopy (ion observed at 1143.5).
- Analytical HPLC Conditions (System F)
- Column: Chromolith RP-18e 4.6 mm×100 mm from EM science
- Gradient: 20-40% B in A over 6 min
- Flow Rate: 4 mL/min
- Solvent A: 0.1% TFA in water
- Solvent B: 0.1% TFA in acetonitrile
- Wavelength: 210 nm, 280 nm
- Preparative HPLC Conditions (System E)
- Column: Waters Delta-Pak radial compression column, 15 um, 100A
- Gradient: 15-25% B in A over 40 min
- Flow Rate: 80 mL/min
- Solvent A: 0.1% acetic acid in water
- Solvent B: acetonitrile
- Wavelength: 214 nm
- Step A: Synthesis of Peptides on Wang Resin
- See Example 1, Step A.
- Step B: Cleavage of Peptides from Wang Resin
- See Example 1, Step B.
- Step C: Coupling of Peptide Acids to 7-Gly-Taxol or 7-Ala-Taxol
- To a mixture of peptide acid (0.0121 mmol, 1.1 equivalents), 7-Gly-Taxol of 7-Ala-Taxol (0.011 mmol), and HATU (0.0154 mmol, 5.9 mg, 1.4 equivalents) was added DMF (0.3 mL) then 2,6-lutidine (0.044 mmol, 5.1 μL, 4 eqiuvalents). The mixture was mixed until a homogeneous solution was obtained. After 4 to 24 hours (monitor by HPLC system H) the reaction was diluted with water (9 mL) and directly purified by preparative HPLC (system 1). Pure fractions were then lyophilized to provide the desired peptide taxol conjugate as a fluffy white solid. The quality of the final conjugate was verified by analytical HPLC (system H) and mass spectroscopy. Typical yields were 30-50% with a purity of 96-99%.
- Analytical HPLC Conditions (System H)
- Column: Chromolith RP-18e 4.6 mm×100 mm from EM science
- Gradient: 5-90% B in A over 6 min
- Flow Rate: 4 mL/min
- Solvent A: 0.1% TFA in water
- Solvent B: 0.1% TFA in acetonitrile
- Wavelength: 210 nm, 280 nm
- Examples of Retention Times (min)
Ac-Gln-Ser-Arg-Ala-Ala-Taxol 2.86 Ac-Gln-Ser-Arg-Ser-Ala-Ala-Taxol 2.79 Ac-Ser-Gly-Arg-Ala-Ser-Ala-Taxol 2.87 Ac-Arg-Ser-Arg-Ala-Ala-Taxol 2.80 Ac-Ser-Gly-Arg-Ser-Ser-Ala-Taxol 2.81 - Preparative HPLC Conditions (System I)
- Column: Ultro 120 5 C18Q 150×20 mm from Peeke Scientific
- Gradient: 20-45% B in A over 40 min
- Flow Rate: 18 mL/min
- Solvent A: 0.1% TFA in water
- Solvent B: acetonitrile
- Wavelength: 214 nm
-
- Step A: N-Ac-Arg-Gln-Ser-Arg-Ala-Ala-OH
- Using the following general procedure, the N-acetyl peptidic substrate N-Ac-Arg-Gln-Ser-Arg-Ala-Ala-OH was synthesized in a peptide synthesis flask. Commencing with commercial Fmoc-Ala-Wang resin (0.35 g, 0.84 mmol, Nova), standard Fmoc-deprotection with 20% piperidine was followed by a sequential iterative coupling-Fmoc deprotection strategy. Each coupling employed a 3-fold excess (2.52 mmol) of Fmoc-Ala, Fmoc-Arg(Boc)2, Fmoc-Ser(tBu), Fmoc-Gln(Trt) and Fmoc-Arg(Boc)2, respectively. Couplings were achieved using PyBOP (2.52 mmol) and DIEA (2.52 mmol) in DMF solvent. During each coupling cycle, the Fmoc protecting group was removed using 20% piperidine in DMF. After removal of the N-terminal Fmoc group, capping with acetic anhydride (1.43 mmol, 1.7 equiv.), DMAP (0.25 mmol, 0.3 equiv.), and DIEA (1.26 mmole, 1.5 equiv.) afforded the resin-bound N-acetyl intermediate. The protected peptide resin was treated with 50% TFA in methylene chloride for 30 min to cleave the Wang resin and then the Boc, Trt and t-Bu protecting groups were removed with 70% TFA in methylene chloride. Solvent and other volatile byproducts were evaporated under reduced pressure and the crude product was dissolved in water and lyophilized to afford the title compound as a nearly colorless, amorphous solid. Mass spectral analysis confirmed the desired molecular weight. HPLC analysis indicated the product to be of approximately 95% purity. The peptide carboxylic acid intermediate can be further purified by trituration or by preparative HPLC, if desired.
- Step B: N-Ac-Arg-Gln-Ser-Arg-Ala-Ala-DOX
- The intermediate from Step A (20 mg, 0.027 mmol) was dissolved in dry DMF (0.8 mL) and was stirred at room temperature under a nitrogen atmosphere. To this solution was added doxorubicin hydrochloride (15.6 mg, 0.027 mmol), EDC (6.8 mg, 0.035 mmol), HOAt (4.8 mg, 0.035 mmol) and 2,6-lutidine (7.3 μL, 0.06 mmol). Stirring was continued until completion of the coupling, as monitored by analytical HPLC (system J, see below). The solution was filtered and the crude product was purified by C18 RP-HPLC (A=0.1% AcOH/H2O; B=CH3CN), gradient elution 100% to 60% A over 60 min). Homogeneous product fractions (evaluated by HPLC, system J) were pooled and lyophilized to afford the title compound as a light red solid.
- HPLC conditions, System J:
- Column: Phenomenex 15 cm #00F-3033-E0, C18
- Eluant: Gradient 95:5 (A:B) to 25:75 (A:B) over 20 min.
- A=0.1% TFA/H2O, B=0.1% TFA/Acetonitrile
- Flow: 1 mL/min.
- Wavelength: 210 nm, 280 nm
- Retention times: Doxorubicin=8.89 min.
- N-Ac-Arg-Gln-Ser-Arg-Ala-Ala-Dox=8.4 min.
- Physical Properties:
- Molecular Formula: C55H78N14O20
- Molecular Weight: 1255.3
- Low Resolution Mass Spec: 628.2 (M+2/2)
- Table 2 lists data for additional peptidic substrate-Doxorubicin conjugates. These conjugates were prepared from the appropriate amino acid precursors that were elaborated by the general procedures described in Example 4.
TABLE 2 HPLC- Mass Retention Peptidic substrate-DOX Conjugate Spectrum Time (min.) Acetyl-Arg-Arg-Gln-Ser-Arg-Ala-Ala-DOX 471.2 8.23 (M + 3/3) Acetyl-Leu-Arg-Arg-Gln-Ser-Arg-Ala-Ala- 509.2 8.60 DOX (M + 3/3) - One millimolar stock solutions were prepared for each peptidic substrate conjugate in double distilled water. Cleavage reactions were then performed in which 100 μM conjugate was mixed with 1 or 10 nM of the recombinantly-produced active single chain protease domain of MTSP1 (residue 615-855 in SEQ ID No. 2, encoded by nucleotides 1865-2582 in SEQ ID No. 1) in 29.2 mM Tris, pH 8.4, 29.2 mM Imidazole, 217 mM NaCl. Final reaction volume was 200 μL. These reactions were incubated in a water bath at 37° C. At times ranging from 2 to 128 minutes, 20 μL samples were removed, and enzymatic activity was stopped by the addition of trifluoroacetic acid to 0.33%. The amount of hydrolysis in each sample was measured by reverse phase HPLC. Percent hydrolysis was then calculated by dividing the area under the product peak by the sum of the areas under substrate and product peaks. Percent unhydrolyzed substrate was plotted against log of reaction times, and the plots were fit to sigmoidal curves using Prism software from Graphpad Inc. (San Diego, Calif.) to determine times at which 50% of each substrate was cleaved.
- Results for certain of the conjugates provided herein are shown in FIG. 1 (conditions: 1 nM MTSP1 with 100 μM conjugate at 37° C. in 12 mM tris(hydroxymethyl)aminomethane, pH 8.0, 25 mM NaCl, 0.5 mM CaCl2; reactions were quenched with 0.33% trifluoroacetic acid).
- The cytotoxicity of the conjugates also can be tested to confirm that the conjugates act as prodrugs. The conjugates are tested against a line of cells, which is known to be killed by unmodified cytotoxic agent, using-an Alamar Blue assay. Cells, such as LNCaP cells (The American Type Culture Collection (Rockville, Md.)), that express a cell surface protease, such as MTSP1 or endotheliase, are seeded in 96 well plates at a density of 1×104 cells/well (0.1 mL/well). A plate containing medium alone is used as a control. The cells are incubated for 3 days at 37° C. and 20 μL of Alamar Blue is added to the assay well(s). After 7 h of incubation, cell killing is measured using an EL-310 plate reader at 570 and 600 nm. Values for cell killing are expressed as the percentage reduction in cell numbers relative to the media controls.
- Tumor cells are trypsinized, resuspended in the growth medium and centrifuged for 6 min at 200×g. The cells are resuspended in serum-free α-MEM and counted. The appropriate volume of this solution containing the desired number of cells is then transferred to a conical centrifuge tube, centrifuged as before and resuspended in the appropriate volume of a cold 1:1 mixture of α-MEM-Matrigel. The suspension is kept on ice until the animals are inoculated.
- Male nude mice 10 weeks of age are used. Mice are individually weighed and assigned to groups (n=10 per group) with no more than a 2-gram difference in weight between individual mice within each group. On
day 1, mice are inoculated subcutaneously with the tumor cell line. ach mouse is inoculated with, for example, 0.5 mL of 0.5×106 to 108 tumor cells/mL in a 60% solution of ice-cold Matrigel and α-MEM. Then, 24 h later, conjugate administration began. Vehicle-treated mice are injected with 5% dextrose in water. At the end of a predetermined time, such as 18 days to two months or more, the mice are sacrificed, and tumor size and mass or other parameters are measured. Tumor size and mass or the other parameters for conjugate-treated mice are compared to vehicle-treated mice to determine efficacy of the conjugate. - Following inoculation with the tumor cells the mice are treated under one of three protocols:
- Protocol A
- One day after cell inoculation the animals are dosed with 1 to 100, or 3 to 50, or 5 to 25, or 7 to 22 μmol/kg, including 7.2 or 17.9 μmol/kg, of test conjugate, unmodified cytotoxic agent or vehicle control (sterile water). Dosages of the conjugate and cytotoxic agent are initially the maximum non-lethal amount, but can be subsequently titrated lower. Identical doses are administered at 24 hour intervals for 5 days. At the end of 5.5 weeks or other suitable interval, the mice are sacrificed and weights of any tumors present are measured. The animals' weights are determined at the beginning and end of the assay.
- Protocol B At 14-15 days after cell inoculation, the animals are dosed with 1 to 100, or 3 to 50, or 5 to 25, or 7 to 22 μmol/kg, including 7.2 or 17.9 μmol/kg, of test conjugate, unmodified cytotoxic agent, or vehicle control (sterile water). Dosages of the conjugate and cytotoxic agent are initially the maximum non-lethal amount, but can be subsequently titrated lower. Identical doses are administered at 24 hour intervals for 5 days. At the end of 5.5 weeks or other suitable interval, the mice are sacrificed and weights of any tumors present are measured. The animals' weights are determined at the beginning and end of the assay.
- Protocol C
- One day after cell inoculation, the animals are dosed by interperitoneal administration with 1 to 100, or 3 to 50, or 5 to 25, or 7 to 22 μmol/kg, including 7.2 or 17.9 μmol/kg, of test conjugate, unmodified cytotoxic agent, or vehicle control (sterile water). Dosages of the conjugate and cytotoxic agent are initially the maximum non-lethal amount, but can be subsequently titrated lower. Identical doses are administered at 7 day intervals for 5 weeks. At the end of 5.5 weeks or other suitable interval, the mice are sacrificed and weights of any tumors present are measured. The animals' weights are determined at the beginning and end of the assay.
- Gene Expression Profile of MTSP1 in Normal Tissues, Cancer Cells and Cancer Tissues
- To obtain information regarding the tissue distribution and gene expression level of MTSP1, the DNA insert from a Pichia pastoris expression vector, pPIC9K-MTSP1, containing the encoding nucleic acid, was used to probe a blot containing RNA from 76 different human tissues (catalog number 7775-1; human multiple tissue expression (MTE) array; CLONTECH, Palo Alto, Calif.). Significant expression was observed in the colon (ascending, transverse and descending), rectum, trachea, esophagus and duodenum. Moderate expression levels were observed in the jejunum, ileum, ilocecum, stomach, prostate, pituitary gland, appendix, kidney, lung, placenta, pancreas, thyroid gland, salivary gland, mammary gland, fetal kidney, and fetal lung. Lower expression levels were seen in the spleen, thymus, peripheral blood leukocyte, lymph node, bone marrow, bladder, uterus, liver, adrenal gland, fetal heart, fetal liver, fetal spleen, and fetal thymus. A significant amount of the MTSP1 transcript was also detected in colorectal adenocarcinoma cell line (SW480), Burkitt's lymphoma cell line (Daudi), and leukemia cell line (HL-60). RT-PCR of the MTSP1 transcript in several human primary tumors xenografted in athymic nude mice was performed using gene-specific primers. A high level of MTSP1 transcript was detected in colon adenocarcinoma (CX-1) and pancreatic adenocarcinoma (GI-103). Moderate levels were observed in another colon adenocarcinoma (GI-112), ovarian carcinoma (GI-102), lung carcinoma (LX-1), and breast carcinoma (GI-101). Another lung carcinoma (GI-117) expressed a low level of the MTSP1 transcript. A similar RT-PCR was performed to detect the presence of the MTSP1 transcript in PC-3 and LNCaP cell lines. Both cell lines expressed significant amounts of MTSP1 transcript. MTSP1 also is a marker for ovarian cancer.
- Gene Expression Profile of the Serine Protease MTSP3 in Normal and Tumor Tissues
- To obtain information regarding the tissue distribution of the MTSP3 transcripts, the DNA insert encoding the MTSP3 protease domain was used to probe a RNA blot composed of 76 different human tissues (catalog number 7775-1; human multiple tissue expression (MTE) array; CLONTECH, Palo Alto, Calif.). The expression pattern observed in decreasing signal level was: trachea=colon (descending)=esophagus>colon (ascending)>colon (transverse)=rectum>ileum>duodenum>jejunum>bladder>ilocecum>stomach>kidney>appendix. It also is expressed less abundantly in fetal kidney, and in two tumor cell lines, HeLa S3 and leukemia, K-562. Northern analysis using RNA blots (catalog numbers 7780-1, 7765-1 & 7782-1; human 12-lane, human muscle and human digestive system multiple tissue northern (MTN) blots; CLONTECH) confirmed that the expression was detected most abundantly in the colon, moderately in the esophagus, small intestine, bladder and kidney, and less abundantly in stomach and rectum. A single transcript of ˜2.2 kb was detected.
- Amplification of the MTSP3 transcript in several human primary tumors xenografted in mouse was performed using gene-specific primers. The MTSP3 transcript was detected in lung carcinoma (LX-1), colon adenocarcinoma (CX-1), colon adenocarcinoma (GI-112) and ovarian carcinoma (GI-102). No apparent signal was detected in another form of lung carcinoma (GI-117), breast carcinoma (GI-101), pancreatic adenocarcinoma (GI-103) and prostatic adenocarcinoma (PC3).
- Gene Expression Profile of MTSP4 in Normal and Tumor Tissues
- To obtain information regarding the gene expression profile of the MTSP4 transcript, a DNA fragment encoding part of the LDL receptor domain and the protease domain was used to probe an RNA blot composed of 76 different human tissues (catalog number 7775-1; human multiple tissue expression (MTE) array; CLONTECH). As in the northern analysis of gel blot, a very strong signal was observed in the liver. Signals in other tissues were observed in (decreasing signal level): fetal liver>heart=kidney=adrenal gland=testis fetal heart and kidney=skeletal muscle=bladder=placenta>brain spinal cord=colon=stomach=spleen=lymph node=bone marrow=trachea=uterus=pancreas=salivary gland=mammary gland=lung. MTSP4 also is expressed less abundantly in several tumor cell lines including HeLa S3=leukemia K-562=Burkitt's lymphomas (Raji and Daudi)=colorectal adenocarcinoma (SW480)>lung carcinoma (A549)=leukemia MOLT-4=leukemia HL-60. PCR of the MTSP4 transcript from cDNA libraries made from several human primary tumors xenografted in nude mice (human tumor multiple tissue cDNA panel, catalog number K1522-1, CLONTECH) was performed using MTSP4-specific primers. The MTSP4 transcript was detected in breast carcinoma (GI-101), lung carcinoma (LX-1), colon adenocarcinoma (GI-112) and pancreatic adenocarcinoma (GI-103). No apparent signal was detected in another form of lung carcinoma (GI-117), colon adenocarcinoma (CX-1), ovarian carcinoma (GI-102). and prostatic adenocarcinoma (PC3). The MTSP4 transcript was also detected in LNCaP and PC-3 prostate cancer cell lines as well as in HT-1080 human fibrosarcoma cell line.
- Gene Expression Profile of MTSP6 in Normal and Tumor Tissues
- To obtain information regarding the gene expression profile of the MTSP6 transcript, a 495 bp DNA fragment obtained from PCR reaction with primers Ch 17-NSP-3 and NSP-4AS was used to probe an RNA blot composed of 76 different human tissues (catalog number 7775-1; human multiple tissue expression (MTE) array; CLONTECH). The strongest signal was observed in duodenum. Signals in other tissues were observed in (decreased signal level): Stomach>trachea=mammary gland=thyroid gland=salivary gland=pituitary gland=pancreas>kidney>lung>jejunum=ileum=ilocecum=appendix=fetal kidney>fetal lung. Very weak signals also can be detected in several other tissues. MTSP6 also is expressed in several tumor cell lines including HeLa S3>colorectal adenocarcinoma (SW480)>leukemia MOLT-4>leukemia K-562. PCR analysis of the MTSP6 transcript from cDNA libraries made from several human primary tumors xenografted in nude mice (human tumor multiple tissue cDNA panel, catalog number K1522-1, CLONTECH) was performed using MTSP6-specific primers (Ch17-NSP-3 and Ch17-NSP2AS). The MTSP6 transcript was strongly detected in lung carcinoma (LX-1), moderately detected in pancreatic adenocarcinoma (GI-103), weakly detected in ovarian carcinoma (GI-102); and very weakly detected in colon adenocarcinoma (GI-112 and CX-1), breast carcinoma (GI-101), lung carcinoma (GI-117) and prostatic adenocarcinoma (PC3). The MTSP6 transcript was also detected in breast cancer cell line MDA-MB-231, prostate cancer cell line PC-3, but not in HT-1080 human fibrosarcoma cell line. MTSP6 also is expressed in mammary gland carcinoma cDNA (Clontech).
- Gene Expression Profile of MTSP9 in Normal, Tumor Tissues and Cell Lines
- To obtain a gene expression profile of the MTSP9 transcript, the MTSP9 cDNA fragment obtained from human pancreas was used to probe a dot blot composed of RNA extracted from 76 different human tissues (Human Multiple Tissue Expression (MTE) Array; Clontech, Palo Alto, Calif.; catalog no. 7775-1). The results of this analysis indicate that MTSP9 is highly expressed in the esophagus and expressed at a low level in many other tissues. The MTSP9 transcript is found in kidney (adult and fetal), spleen (adult and fetal), placenta, liver (adult and fetal), thymus, peripheral blood leukocyte, lung (adult and fetal), pancreas, lymph node, bone marrow, trachea, uterus, prostate, esophagus, testes, ovary and the gland organs (mammary, adrenal, thyroid, pituitary and salivary). MTSP9 also is expressed in tumor esophagus tissues, in a lung carcinoma (A549 cell line) and, at a low level, in a colorectal carcinoma (SW480), lymphoma (Raji and Daudi), a cervical carcinoma (HeLaS3) and leukemia (HL-60, K-562 and MOLT-4) cell lines.
- Gene Expression Profile of MTSP10 in Normal and Tumor Tissues
- To obtain information regarding the gene expression profile of the MTSP10 transcript, PCR analysis was carried out on cDNA panels made from several human adult tissues (Clontech, Cat. #K1420-1) cDNA panel using MTSP10-specific primers. MTSP10 transcript was detected in pancreas, lung and kidney. MTSP10 transcript was also detected in small intestine Marathon-Ready cDNA (Clontech). PCR of the MTSP10 transcript from cDNA libraries made from several human primary tumors xenografted in nude mice (human tumor multiple tissue cDNA panel, catalog number K1522-1, CLONTECH) was also performed. The MTSP10 transcript was detected in breast carcinoma (GI-101), lung carcinoma (LX-1 and GI-117), ovarian carcinoma (GI-102), and pancreatic adenocarcinoma (GI-103). The MTSP10 transcript can be weakly detected in prostatic adenocarcinoma (PC3). No apparent signal was detected in two forms of colon adenocarcinomas (GI-112 and CX-1). The MTSP10 transcript was also detected in CWR22R prostate tumor grown on nude mice.
- Domain Organization and Gene Expression Profile of MTSP12 in Normal and Tumor Tissues
- Domain Organization of MTSP12PD1, -PD2 and -PD3 and Homology to other Serine Proteases
- Sequence and protein domain analyses of the translated MTSP12PD1, -PD2 and -PD3 nucleotide sequences indicate that these three serine proteases are contiguous. The sequence order is as follows: MTSP12-PD1 is found at the N terminus followed by MTSP12-PD2, and MTSP12-PD3 is at the C terminus. MTSP12-PD1 and -PD2 contain a trypsin-like serine protease domain (aa 236 to
aa 465 andaa 537 to aa 765 for MTSP12-PD1 and -PD2, respectively) characterized by the presence of a protease activation cleavage site ( . . . R236↓I237VGGMEAS . . . , and . . . R537↓V538VGGFGAA . . . , for MTSP12-PD1 and -PD2, respectively, and where ¦ indicates a protease activation cleavage site) and the catalytic triad residues (His277, Asp326 and Ser421 in MTSP12-PD1; His578, Asp626 and Ser721 in MTSP12-PD2) in 3 highly-conserved regions of the catalytic domain. MTSP12-PD3 contains a serine protease domain (aa 861 to aa 1087); it has a protease activation cleavage site ( . . . R860↓I861VGGSAAG . . . ) and has the catalytic His902 and Asp949, but it has a Ala1043 instead of the conserved catalytic serine found in serine proteases. Several domains are found upstream of the MTSP12-PD1 serine protease domain and these include a transmembrane domain (aa 28 to aa 50), a SEA (sea urchin sperm protein-enterokinase-agrin) domain (aa 51 to aa 170) and an LDLa (low density lipoprotein receptor class a) domain (aa 187 to aa 225). There are 5 possible N-linked glycosylation sites (N116SS, N581HT, N672AT, N697FS and N820ST). In the protease domain of MTSP12-PD1, there is an unpaired cysteine (C346) in a single chain form of the protease domain and the following Cys pairings are noted: C262-C278; C360-C427; C417-C446; C392-C406. In the protease domain of MTSP12-PD2, there is an unpaired cysteine (C646) in a single chain form of the protease domain, and the following Cys pairings are noted: C563-C579; C660-C727; C692-C706; C717-C746. In the protease domain of MTSP12-PD3, there is an unpaired cysteine (C969) in a single chain form of the protease domain, and the following Cys pairings are noted: C887-C903; C983-C1049; C1014-C1028; C1039-C1068. - Alignment (blastp; http://www.ncbi.nlm.nih.gov/BLAST) of the respective MTSP12-PD1, MTSP12-PD2 and MTSP12-PD3 protein sequences to known serine proteases deposited in the public database showed a 45%, 45% and 48% identity to matriptase, a 44%, 43% and 41% identity with DESC1/
endotheliase 1, a 44%, 43% and 48% identity to prostamin (AB030036), a 43%, 39% and 39% identity to spinesin (TMPRSS5; NM—030770), and a 40%, 38% and 38% identity to marapsin (NM—031948). The clone has about 93% homology at the nucleotide and encoded protein levels to a clone and encoded provided described in International PCT application No. WO 02/00860 (see SEQ ID Nos. 38 and 97 therein). The encoded protein described in the PCT application, however, includes the Sequence set forth in SEQ ID No. 271 between amino acids Leu373 and Val374 of SEQ ID No. 20, as well as an additional extended sequence of amino acids beteween amino acids Ala48 and Phe49 of SEQ ID No. 20 and lacks amino acids 91-124 of SEQ ID No. 20. The protein provided in International PCT application No. WO02/00860 can be used in the methods provided herein. - Gene and Tissue Expression Profile of MTSP12
- To obtain information regarding the tissue distribution profile of the MTSP12PD1, -PD2 and -PD3 transcripts, 3 cDNA probes were prepared. Data indicate that the MTSP12PD1, -PD2 and -PD3 transcript is expressed at a low level in most of the 76 tissues and cell lines, but at a higher level in the lymph node and testes.
- To compare the expression profile of MTSP12PD1, -PD2 and -PD3 in a range of normal human and matched tumor tissues, a matched tumor/normal expression array (catalog number 7840-1; http://www.clontech.com) composed of 68 paired cDNA samples from individual patients was used. Results show that the MTSP12PD1, -PD2 and -PD3 transcript is expressed at a low level in a number of normal tissues including breast, uterus, colon, ovary, lung, kidney and rectum, but is not differentially expressed in any of the matched tumors. It also is expressed at a low level in several tumor cell lines, including HeLa (cervical carcinoma), Daudi (Burkitt's lymphoma), K562 (chronic myelogenous leukemia), HL-60 (premyelocytic leukemia), G361 (melanoma), A549 (lung carcinoma), MOLT-4 (lymphoblastic leukemia), SW480 (colorectal adenocarcinoma), and Raji (Burkitt's lymphoma).
- Several SMART™ 5′-RACE cDNA libraries (catalog number K1811-1; http://www.clontech.com) prepared from normal breast, normal testes, normal prostate, prostate cancer cell lines and breast cancer cell lines were analyzed for the presence of MTSP12PD1, -PD2 and -PD3 transcript by RT-PCR using two sets of gene-specific primers. The MTSP12-PD2 and -PD3 transcript was detected in normal prostate, PC-3, LNCaP, normal breast, MDA-MB-231, MDA-MB-361, MDA-MB-453 and DU4475, but higher levels were observed in normal breast and MDA-MB-231. The MTSP12-PD1 transcript was detected in the same tissues and cell lines, except higher levels were observed in normal breast, MDA-MB-231 and DU4475.
- Gene Expression Profile of MTSP20 in Normal, Tumor Tissues and Cell Lines
- To obtain information regarding the gene expression profile of the MTSP20 transcript, the MTSP20 cDNA fragment obtained from human lung tissue was used to probe a dot blot composed of RNA extracted from 76 different human tissues (Human Multiple Tissue Expression (MTE) Array; Clontech, Palo Alto, Calif.; catalog no. 7775-1). The results indicate that RNA encoding MTSP20 is expressed in a variety of tissues. The MTSP20 transcript is found in liver, lymph node, cerebellum, pancreas, prostate, uterus, testis, glands (adrenal, thyroid and salivary), thymus, kidney and spleen. Lower transcript level can be found in lung, placenta, bladder, ovary, digestive system, circulatory system and other parts of the the brain. MTSP20 is also expressed in certain tumor cell lines including lung carcinoma (A519), colorectal carcinoma (SW480), lymphoma (Raji and Daudi), cervical carcinoma (HeLaS3) and leukemia (HL-60, K-562 and MOLT-4) cell lines.
- Gene Expression Profile of MTSP22 in Normal, Tumor Tissues and Cell Lines
- MTSP22 is expressed in the uterine tissue, thymus, adipose tissue, and lymph node. It may also be expressed in lung, stomach, uterine, breast, ovarian, prostate and in other tumors. To obtain information regarding the gene expression profile of the MTSP22 transcript, the cDNA fragment encoding the entire serine protease domain was used to probe a dot blot composed of RNA extracted from 72 different human tissues (Human Multiple Tissue Expression (MTE) Array; Clontech, Palo Alto, Calif.; catalog no. 7776-1) as well as a dot blot composed of normalized cDNA from 241 tumor and corresponding normal tissues from individual patients (Cancer Profiling Array, Clontech, catalog no. 7841-1). The results of MTE analysis indicated that MTSP22 transcript is expressed primarily in the esophagus. In the cancer profiling array analysis, MTSP22 is highly expressed in 3 of the 42 normal uterus tissue samples, but not in their matched tumor samples. In one of the 42 uterus samples, MTSP22 is expressed in tumor and its metastatic tissues, but not in the normal tissue counterpart. MTSP22 is also expressed in 2 of the 17 stomach tumors and 2 of the 21 lung tumors, but not in their normal tissue counterparts. MTSP22 is also expressed in the normal tissue of the only pancreas matched cDNA pair. PCR analysis was also performed using commercially available cDNA panel from several human adult tissues (Clontech, Cat. #K1420-1 and K1420-2) and primary tumors (Clontech Cat. #K1522-1) as well as several Marathon-Ready cDNAs (Clontech).
- MTSP22 cDNA was detected in thymus, adipose tissue, and lymph node. Serine protease domain of MTSP22 and homology to other proteases.
- Sequence analysis of the translated MTSP22 protease domain sequence revealed that MTSP22 contains a trypsin-like serine protease domain characterized by the presence of a protease activation cleavage site at the amino terminus of the domain and the catalytic triad residues (histidine, aspartate and serine) in three highly-conserved regions. Alignment of the protein sequence with that of endotheliase 1 (same as serine protease DESC1 protein; GenBank accession number AF064819) indicated that the two proteins share 50% sequence identity in their protease domains.
- Gene Expression Profile of MTSP25 in Normal, Tumor Tissues and Cell Lines
- MTSP25 is expressed in breast, colon, uterine, ovarian, kidney, prostate, testicular cancer tissue. It may also be expressed in lung, stomach, prostate and in other tumors. To obtain information regarding the gene expression profile of the MTSP25 transcript, a 369 bp DNA fragment containing MTSP25 protease domain sequence (obtained from a PCR reaction) was used to probe a dot blot composed of RNA extracted from 72 different human tissues (Human Multiple Tissue Expression (MTE) Array; Clontech, Palo Alto, Calif.; catalog no. 7776-1) as well as a dot blot composed of normalized cDNA from 241 tumor and corresponding normal tissues from individual patients (Cancer Profiling Array, Clontech, catalog no. 7841-1). The results of MTE analysis indicate that MTSP25 transcript is expressed weakly in the lymph node. In the cancer profiling array analysis, MTSP25 is highly expressed in all 4 prostate samples (in normal and cancer samples). In one of the 20 kidney cDNA pairs, MTSP25 is highly expressed in the tumor sample, but not in its normal tissue counterpart. MTSP25 is also expressed in 1 of the 50 breast cancer samples, but not in its normal tissue counterpart.
- MTSP25 is also expressed in 3 of the 42 normal uterus samples, but not in their tumor counterparts. MTSP25 expression is also detected in 3 of the 14 ovarian cancer samples. Among these three samples, the expression of MTSP25 was also detected in one of the matched normal tissue counterparts. MTSP25 expression was also detected in 5 tumor samples in the 34 colon cDNA pairs.
- PCR analysis was also performed using a commercially available cDNA panel from several human adult tissues (Clontech, Cat. #K1420-1 and K1420-2) as well as several Marathon-Ready cDNAs (Clontech). MTSP25 cDNA was strongly detected in testis and mammary gland adenocarcinoma, weakly detected in brain, placenta, lung, spleen, prostate, small intestine, colon, and leukocyte, and very weakly detected in heart, liver, and pancreas.
- Conjugates that have been prepared according to the procedures of Examples 1-4 by routine and minor modification of the procedures, such as using different Fmoc-amino acid building blocks, include:
- Ac-R-Q-G-R-S-L-(Dox) (SEQ ID NO: 491);
- Ac-R-Q-G-R-S-S-L-(Dox) (SEQ ID NO: 492);
- Ac-R-Q-G-R-S-nL-(Dox) (SEQ ID NO: 493);
- Ac-R-Q-G-R-S-nV-(Dox) (SEQ ID NO: 494);
- Ac-R-Q-G-R-S-F-(Dox) (SEQ ID NO: 495);
- Ac-R-Q-G-R-A-L-(Dox) (SEQ ID NO: 496);
- Ac-R-Q-G-R-A-L-(Dox) (SEQ ID NO: 497);
- Ac-R-Q-G-R-A-nL-(Dox) (SEQ ID NO: 498);
- Ac-R-Q-G-R-A-nL-(Dox) (SEQ ID NO: 499);
- Ac-R-Q-G-R-A-nV-(Dox) (SEQ ID NO: 500);
- Ac-R-Q-G-R-A-Cha-(Dox) (SEQ ID NO: 501);
- Ac-R-Q-G-R-A-F-(Dox) (SEQ ID NO: 502);
- Ac-R-N-G-R-S-L-(Dox) (SEQ ID NO: 503);
- Ac-R-N-G-R-A-nL-(Dox) (SEQ ID NO: 504);
- Ac-R-Q-A-R-S-L-(Dox) (SEQ ID NO: 505);
- Ac-R-Q-A-R-S-nL-(Dox) (SEQ ID NO: 506);
- Ac-R-Q-A-R-S-nV-(Dox) (SEQ ID NO: 507);
- Ac-R-Q-A-A-S-Cha-(Dox) (SEQ ID NO: 508);
- Ac-R-Q-A-R-S-S-Cha-(Dox) (SEQ ID NO: 509);
- Ac-R-Q-A-R-T-nL-(Dox) (SEQ ID NO: 510);
- Ac-R-Q-A-R-A-L-(Dox) (SEQ ID NO: 511);
- Ac-R-Q-A-R-A-nL-(Dox) (SEQ ID NO: 513);
- Ac-R-Q-A-R-A-nV-(Dox) (SEQ ID NO: 514);
- Ac-R-Q-A-R-A-Cha-(Dox) (SEQ ID NO: 515);
- Ac-R-Q-S-R-A-A-(Dox) (SEQ ID NO: 516);
- Ac-R-Q-S-R-A-(Dox) (SEQ ID NO: 517);
- Ac-R-Q-S-R-A-nL-(Dox) (SEQ ID NO: 518);
- Ac-R-Q-S-R-A-L-(Dox) (SEQ ID NO: 519);
- Ac-R-Q-S-R-A-nV-(Dox) (SEQ ID NO: 520);
- Ac-R-Q-S-R-A-Cha-(Dox) (SEQ ID NO: 521);
- Ac-R-Q-S-R-S-S-L-(Dox) (SEQ ID NO: 522);
- Ac-R-Q-S-R-S-L-(Dox) (SEQ ID NO: 523);
- Ac-R-Q-S-R-S-dnL-(Dox) (SEQ ID NO: 524);
- Ac-R-Q-S-R-S-nL-(Dox) (SEQ ID NO: 525);
- Ac-R-Q-S-R-S-nV-(Dox) (SEQ ID NO: 526);
- Ac-R-Q-S-R-S-allylG-(Dox) (SEQ ID NO: 527);
- Ac-R-Q-S-R-S-Cha-(Dox) (SEQ ID NO: 528);
- Ac-R-Q-S-R-T-nL-(Dox) (SEQ ID NO: 529);
- Ac-R-Q-T-R-S-S-L-(Dox) (SEQ ID NO: 530);
- Ac-R-Q-T-R-S-L-(Dox) (SEQ ID NO: 531);
- Ac-R-N-S-R-S-nL-(Dox) (SEQ ID NO: 532);
- Ac-R-Q-F-R-S-L-(Dox) (SEQ ID NO: 533);
- Ac-R-Q-F-R-S-nL-(Dox) (SEQ ID NO: 534);
- Ac-R-Q-F-R-S-nV-(Dox) (SEQ ID NO: 535);
- Ac-R-Q-F-R-S-nL-(Dox) (SEQ ID NO: 536);
- Ac-R-Q-F-R-S-Cha-(Dox) (SEQ ID NO: 537);
- Ac-R-Q-F-R-A-L-(Dox) (SEQ ID NO: 538);
- Ac-R-Q-F-R-A-nL-(Dox) (SEQ ID NO: 539);
- Ac-R-Q-F-R-A-nV-(Dox) (SEQ ID NO: 540);
- Ac-R-Q-F-R-A-Cha-(Dox) (SEQ ID NO: 541);
- Ac-Q-S-R-S-S-nL-(Dox) (SEQ ID NO: 542);
- MeOCO-Quat2-G-R-S-L-NH2 (SEQ ID NO: 483);
- MeOCO-Quat3-G-R-S-L-NH2 (SEQ ID NO: 484);
- MeOCO-Quat-G-R-S-L-NH2 (SEQ ID NO: 485);
- MeOCO-Quat4-G-R-S-L-NH2 (SEQ ID NO: 486);
- MeOCO-Quat5-G-R-S-L-NH2 (SEQ ID NO: 487);
- MeOCO-Quat2-G-R-S-S-L-NH2 (SEQ ID NO: 488);
- MeOCO-Quat4-G-R-S-L-(Dox) (SEQ ID NO: 489);
- MeOCO-Quat2-G-R-S-L-(Dox) (SEQ ID NO: 490);
- Ac-Q-G-R-S-L-(Dox) (SEQ ID NO: 445);
- Ac-Q-G-R-S-S-L-(Dox) (SEQ ID NO: 446);
- Ac-Q-G-R-A-S-L-(Dox) (SEQ ID NO: 447);
- Ac-N-G-R-S-S-L-(Dox) (SEQ ID NO: 448);
- Ac-Q-G-R-S-S-nL-(Dox) (SEQ ID NO: 449);
- Ac-Q-G-R-S-S-nV-(Dox) (SEQ ID NO: 450);
- Ac-Q-G-R-S-S-Cha-(Dox) (SEQ ID NO: 451);
- Ac-Q-G-R-S-S-allylG-(Dox) (SEQ ID NO: 452);
- Ac-Q-G-R-S-S-allylG-(Dox) (SEQ ID NO: 453);
- Ac-Q-A-R-S-L-(Dox) (SEQ ID NO: 454);
- Ac-Q-A-R-S-S-L-(Dox) (SEQ ID NO: 455);
- Ac-Q-S-R-S-L-(Dox) (SEQ ID NO: 456);
- Ac-Q-S-R-S-S-nV-(Dox) (SEQ ID NO: 457);
- Ac-Q-S-R-S-S-Cha-(Dox) (SEQ ID NO: 458);
- Ac-Q-S-R-S-S-L-(Dox) (SEQ ID NO: 459);
- Ac-Q-T-R-S-S-L-(Dox) (SEQ ID NO: 460);
- Ac-Q-Aib-R-S-S-Cha-(Dox) (SEQ ID NO: 461);
- Ac-Q-Aib-R-S-S-L-(Dox) (SEQ ID NO: 462);
- Ac-Q-Abu-R-S-S-Cha-(Dox) (SEQ ID NO: 463);
- Ac-Q-Abu-R-S-S-L-(Dox) (SEQ ID NO: 464);
- Ac-Q-Cha-R-S-S-Cha-(Dox) (SEQ ID NO: 465);
- Ac-Q-F-R-S-L-(Dox) (SEQ ID NO: 466);
- Ac-Q-F-R-S-S-L-(Dox) (SEQ ID NO: 467);
- Ac-Q-Y-R-S-S-L-(Dox) (SEQ ID NO: 468);
- Ac-R-G-R-S-L-(Dox) (SEQ ID NO: 469);
- Ac-R-G-R-S-S-L-(Dox) (SEQ ID NO: 470);
- Ac-R-G-R-S-S-Cha-(Dox) (SEQ ID NO: 471);
- Ac-R-G-R-S-Cha-(Dox) (SEQ ID NO: 472);
- Ac-R-A-R-S-L-(Dox) (SEQ ID NO: 473);
- Ac-R-A-R-S-S-L-(Dox) (SEQ ID NO: 474);
- Ac-R-S-R-S-L-(Dox) (SEQ ID NO: 475);
- Ac-R-S-R-S-S-L-(Dox) (SEQ ID NO: 476);
- Ac-R-S-R-S-Cha-(Dox) (SEQ ID NO: 477);
- Ac-R-S-R-S-S-Cha-(Dox) (SEQ ID NO: 478);
- Ac-R-F-R-S-L-(Dox) (SEQ ID NO: 479);
- Ac-R-F-R-S-Cha-(Dox) (SEQ ID NO: 480);
- Ac-Y-G-R-S-S-L-(Dox) (SEQ ID NO: 481);
- Ac-M(O2)-S-R-S-L-(Dox) (SEQ ID NO: 482);
- Ac-R-R-Q-S-R-A-A-(Dox) (SEQ ID NO: 105);
- Ac-R-R-Q-S-R-I-(Dox) (SEQ ID NO: 610);
- Ac-R-R-Q-S-R-S-S-L-(Dox) (SEQ ID NO: 543);
- Ac-R-R-Q-S-R-S-L-(Dox) (SEQ ID NO: 544);
- Ac-R-G-S-G-R-S-L-(Dox) (SEQ ID NO: 545);
- Ac-R-G-S-G-R--S-nL-(Dox) (SEQ ID NO: 546);
- Ac-R-G-S-G-R-A-nL-(Dox) (SEQ ID NO: 547);
- Ac-R-G-S-G-R-S-S-L-(Dox) (SEQ ID NO: 548);
- Ac-I-V-S-G-R-A-S-L-(Dox) (SEQ ID NO: 549);
- Ac-R-R-Q-S-R-A-(Dox) (SEQ ID NO: 108);
- Ac-R-R-Q-S-R-I-(Dox) (SEQ ID NO: 111);
- Ac-L-R-R-Q-S-R-A-A-(Dox) (SEQ ID NO: 106);
- Ac-L-R-R-Q-S-R-G-G-(Dox) (SEQ ID NO: 109);
- Ac-L-R-R-Q-S-R-A-(Dox) (SEQ ID NO: 110);
- Ac-L-R-R-Q-S-R-A-I-(Dox) (SEQ ID NO: 112);
- Ac-L-R-R-Q-S-R-A-I-(Dox) (SEQ ID NO: 611);
- Ac-L-R-R-Q-S-R-S-S-L-(Dox) (SEQ ID NO: 550);
- Ac-L-R-R-Q-S-R-S-L-(Dox) (SEQ ID NO: 551);
- Ac-S-G-R-S-L-(Dox) (SEQ ID NO: 362);
- Ac-S-G-R-S-S-L-(Dox) (SEQ ID NO: 363);
- Ac-S-G-R-S-S-S-L-(Dox) (SEQ ID NO: 364);
- Ac-S-G-R-S-nL-(Dox) (SEQ ID NO: 365);
- Ac-S-G-R-S-nV-(Dox) (SEQ ID NO: 366);
isomer 1 - Ac-S-G-R-S-nV-(Dox) (SEQ ID NO: 367);
isomer 2 - Ac-S-G-R-S-G(hex)-(Dox) (SEQ ID NO: 368);
- Ac-S-G-R-S-Cha-(Dox) (SEQ ID NO: 369);
- Ac-S-G-R-S-hCha-(Dox) (SEQ ID NO: 370);
- Ac-S-A-R-S-L-(Dox) (SEQ ID NO: 371);
- Ac-S-A-R-S-S-L-(Dox) (SEQ ID NO: 372);
- Ac-S-S-R-S-nL-(Dox) (SEQ ID NO: 373);
- Ac-T-G-R-S-Abu-(Dox) (SEQ ID NO: 374);
- Ac-T-G-R-S-L-(Dox) (SEQ ID NO: 375);
- Ac-T-G-R-S-nV-(Dox) (SEQ ID NO: 376);
- Ac-T-G-R-S-nL-(Dox) (SEQ ID NO: 377);
- Ac-T-G-R-S-G(hex)-(Dox) (SEQ ID NO: 378);
- Ac-T-G-R-S-Cha-(Dox) (SEQ ID NO: 379);
- Ac-T-G-R-S-hCha-(Dox) (SEQ ID NO: 380);
- Ac-T-G-R-T-Abu-(Dox) (SEQ ID NO: 381);
- Ac-T-G-R-hS-nL-(Dox) (SEQ ID NO: 382);
- Ac-T-G-R-Abu-nL-(Dox) (SEQ ID NO: 383);
- Ac-T-G-R-Abu-nV-(Dox) (SEQ ID NO: 384);
- Ac-T-G-F(Gn)-S-nL-(Dox) (SEQ ID NO: 385);
- Ac-T-G-F(Gn)-S-Cha-(Dox) (SEQ ID NO: 386);
- Ac-T-G-F(Gn)-Abu-nV-(Dox) (SEQ ID NO: 387);
- Ac-T-G-K(alloc)-S-nL-(Dox) (SEQ ID NO: 388);
- Ac-T-G-K-S-nL-(Dox) (SEQ ID NO: 389);
- Ac-T-G-hR-S-nL-(Dox) (SEQ ID NO: 390);
- Ac-(hS)G-G-R-S-nL-(Dox) (SEQ ID NO: 391);
- MeOCO-T-G-R-S-nL-(Dox) (SEQ ID NO: 392);
- PhSO2-T-G-R-S-nL-(Dox) (SEQ ID NO: 393);
- MeOEtCO-T-G-R-S-nL-(Dox) (SEQ ID NO: 394);
- MeO(EtO)2Ac-T-G-R-S-nL-(Dox) (SEQ ID NO: 395);
- 4-oxo-Pentanoyl-T-G-R-S-nL-(Dox) (SEQ ID NO: 396);
- 3,4-MethyldioxyPhAc-T-G-R-S-nL-(Dox) (SEQ ID NO: 397);
- 2-PyridilAc-T-G-R-S-nL-(Dox) (SEQ ID NO: 398);
- PhOAc-T-G-R-S-nL-(Dox) (SEQ ID NO: 399);
- L-3-PhLactyl-T-G-R-S-nL-(Dox) (SEQ ID NO: 400);
- MeOAc-T-G-R-S-nL-(Dox) (SEQ ID NO: 401);
- PhAc-T-G-R-S-nL-(Dox) (SEQ ID NO: 402);
- MeOEtOCO-T-G-R-S-nL-(Dox) (SEQ ID NO: 403);
- MeOEtOAc-T-G-R-S-nL-(Dox) (SEQ ID NO: 404);
- HOOCButa-T-G-R-S-nL-(Dox) (SEQ ID NO: 405);
- Z-T-G-R-S-nL-(Dox) (SEQ ID NO: 406);
- EtOCO-T-G-R-S-nL-(Dox) (SEQ ID NO: 407);
- βA-T-G-R-S-nL-(Dox) (SEQ ID NO: 408);
- Pent-4-ynoyl-T-G-R-S-nL-(Dox) (SEQ ID NO: 409);
- NapAc-T-G-R-S-nL-(Dox) (SEQ ID NO: 410);
- iBoc-T-G-R-S-nL-(Dox) (SEQ ID NO: 411);
- HOAc-T-G-R-S-nL-(Dox) (SEQ ID NO: 412);
- MeSucc-T-G-R-S-nL-(Dox) (SEQ ID NO: 413);
- N,N-diMeGly-T-G-R-S-nL-(Dox) (SEQ ID NO: 414);
- Succ-T-G-R-S-nL-(Dox) (SEQ ID NO: 415);
- HCO-T-G-R-S-nL-(Dox) (SEQ ID NO: 416);
- Ac-T-A-R-S-nL-(Dox) (SEQ ID NO: 417);
- Ac-T-A-F(Gn)-S-nL-(Dox) (SEQ ID NO: 418);
- Ac-T-A-R-Abu-nV-(Dox) (SEQ ID NO: 419);
- Ac-T-A-R-S-Abu-(Dox) (SEQ ID NO: 420);
- Ac-T-A-R-T-Abu-(Dox) (SEQ ID NO: 421);
- Ac-T-S(O-Me)-R-S-nL-(Dox) (SEQ ID NO: 422);
- Ac-T-hS-R-S-nL-(Dox) (SEQ ID NO: 423);
- Ac-T-(1-Me)H-R-S-nL-(Dox) (SEQ ID NO: 424);
- Ac-T-(3-Me)H-R-S-nL-(Dox) (SEQ ID NO: 425);
- Ac-T-H-R-S-nL-(Dox) (SEQ ID NO: 426);
- Ac-T-Sar-R-S-nL-(Dox) (SEQ ID NO: 427);
- Ac-T-nV-R-S-nL-(Dox) (SEQ ID NO: 428);
- Ac-T-nL-R-S-nL-(Dox) (SEQ ID NO: 429);
- Ac-T-A-R-S-Cha-(Dox) (SEQ ID NO: 430);
- Ac-T-Abu-R-S-nL-(Dox) (SEQ ID NO: 431);
- Ac-4,4diMeThr-G-R-S-nL-(Dox) (SEQ ID NO: 432);
- Ac-hS-G-R-S-nL-(Dox) (SEQ ID NO: 433);
- Ac-hS-G-R-hS-Cha-(Dox) (SEQ ID NO: 434);
- Ac-hS-G-R-S-Cha-(Dox) (SEQ ID NO: 435);
- Ac-hS-G-R-T-Cha-(Dox) (SEQ ID NO: 436);
- Ac-hS-A-R-S-Cha-(Dox) (SEQ ID NO: 437);
- Ac-N-G-R-S-nL-(Dox) (SEQ ID NO: 438);
- Ac-Y-G-R-S-S-L-(Dox) (SEQ ID NO: 439);
- Ac-Y-G-R-S-Cha-(Dox) (SEQ ID NO: 440);
- Ac-Q-G-R-S-S-nL-(Dox) (SEQ ID NO: 441);
- Ac-Q-G-R-S-S-nV-(Dox) (SEQ ID NO: 442);
- Ac-L-R-G-S-G-R-S-A-(Dox) (SEQ ID NO: 573);
- Ac-L-R-G-S-G-R-S-L-(Dox) (SEQ ID NO: 342);
- Ac-L-R-G-S-G-R-S-L-(Dox) (SEQ ID NO: 343);
- Ac-L-R-G-S-G-R-S-S-nL-(Dox) (SEQ ID NO: 344);
- Ac-L-R-G-S-G-R-S-S-Cha-(Dox) (SEQ ID NO: 345);
- Ac-L-R-G-dS-A-R-S-A-(Dox) (SEQ ID NO: 574);
- Ac-L-R-G-S-A-R-S-S-L-(Dox) (SEQ ID NO: 346);
- Ac-L-R-G-S-A-R-S-L-(Dox) (SEQ ID NO: 347);
- Ac-L-R-G-S-A-R-S-S-Cha-(Dox) (SEQ ID NO: 348);
- Ac-L-R-G-S-A-R-S-S-nV-(Dox) (SEQ ID NO: 349);
- Ac-L-R-G-S-A-R-S-S-nL-(Dox) (SEQ ID NO: 350);
- Ac-V-I-V-S-G-R-A-L-(Dox) (SEQ ID NO: 351);
- Ac-V-I-V-S-A-R-S-L-(Dox) (SEQ ID NO: 352);
- Ac-V-I-V-S-G-R-S-S-L-(Dox) (SEQ ID NO: 353);
- Ac-V-I-V-S-A-R-M-A-(Dox) (SEQ ID NO: 354);
- Ac-V-I-V-S-A-R-nL-A-(Dox) (SEQ ID NO: 355);
- Ac-V-I-V-S-A-R-S-nL-(Dox) (SEQ ID NO: 356);
- Ac-V-I-V-S-A-R-S-Cha-(Dox) (SEQ ID NO: 357);
- Ac-V-I-V-S-A-R-S-Cha-(Dox) (SEQ ID NO: 358);
- Ac-V-I-V-S-A-R-S-S-Cha-(Dox) (SEQ ID NO: 359);
- Ac-R-R-(Me)C-P-G-R-V-V-(Dox) (SEQ ID NO: 360);
- Ac-R-R-nV-P-A-R-S-L-(Dox) (SEQ ID NO: 361);
- Ac-R-G-dS-A-R-S-A-(Dox) (SEQ ID NO: 309);
- Ac-R-G-S-G-R-S-A-(Dox) (SEQ ID NO: 310);
- Ac-R-G-S-G-R-A-L-(Dox) (SEQ ID NO: 311);
- Ac-R-G-S-G-R-S-L-(Dox) (SEQ ID NO: 312);
- Ac-R-G-S-G-R--S-nL-(Dox) (SEQ ID NO: 313);
- Ac-R-G-S-G-R-A-nL-(Dox) (SEQ ID NO: 314);
- Ac-R-G-S-G-R-S-S-L-(Dox) (SEQ ID NO: 315);
- Ac-R-G-S-G-R-S-Cha-(Dox) (SEQ ID NO: 316);
- Ac-R-G-S-G-R-S-S-Cha-(Dox) (SEQ ID NO: 317);
- Ac-R-G-S-A-R-S-Cha-(Dox) (SEQ ID NO: 318);
- Ac-R-G-S-A-R-S-S-(Dox) (SEQ ID NO: 319);
- Ac-R-G-S-A-R-S-nV-(Dox) (SEQ ID NO: 320);
- Ac-R-G-S-A-R-S-S-nV-(Dox) (SEQ ID NO: 321);
- Ac-R-G-S-A-R-S-L-(Dox) (SEQ ID NO: 322);
- Ac-R-(Me)C-P-G-R-V-V-(Dox) (SEQ ID NO: 323);
- Ac-R-(Me)C-P-G-R-V-V-(Dox) (SEQ ID NO: 324);
- Ac-R-C(Me)-P-G-R-S-L-(Dox) (SEQ ID NO: 325);
- Ac-R-L-P-G-R-S-L-(Dox) (SEQ ID NO: 326);
- Ac-R-V-P-G-R-S-L-(Dox) (SEQ ID NO: 327);
- Ac-R-V-P-G-R-S-L-(Dox) (SEQ ID NO: 328);
- Ac-R-nL-P-G-R-S-L-(Dox) (SEQ ID NO: 329);
- Ac-R-G(tBu)-P-A-R-S-L-(Dox) (SEQ ID NO: 330);
- Ac-R-L-P-A-R-S-L-(Dox) (SEQ ID NO: 331);
- Ac-R-V-P-A-R-S-L-(Dox) (SEQ ID NO: 332);
- Ac-R-nL-P-A-R-S-L-(Dox) (SEQ ID NO: 333);
- Ac-I-V-S-G-R-A-L-(Dox) (SEQ ID NO: 334);
- Ac-I-V-S-G-R-S-S-L-(Dox) (SEQ ID NO: 335);
- Ac-I-V-S-G-R-A-S-L-(Dox) (SEQ ID NO: 336);
- Ac-I-V-S-A-R-M-A-(Dox) (SEQ ID NO: 337);
- Ac-I-V-S-A-R-nL-A-(Dox) (SEQ ID NO: 338);
- Ac-I-V-S-A-R-S-L-(Dox) (SEQ ID NO: 339);
- Ac-I-V-S-A-R-S-nL-(Dox) (SEQ ID NO: 340);
- Ac-I-V-S-A-R-S-S-L-(Dox) (SEQ ID NO: 341);
- Ac-G-S-G-R-S-A-(Dox) (SEQ ID NO: 585);
- Ac-G-S-G-R-S-L-(Dox) (SEQ ID NO: 277);
- Ac-G-S-G-R-A-L-(Dox) (SEQ ID NO: 278);
- Ac-G-S-G-R-S-S-L-(Dox) (SEQ ID NO: 279);
- Ac-G-S-G-R-L-(Dox) (SEQ ID NO: 280);
- Ac-G-S-G-(4-guan)Phg-S-L-NH2 (SEQ ID NO: 281);
- Ac-G-S-G-R-S-S-Cha-(Dox) (SEQ ID NO: 282);
- Ac-G-S-G-R-A-S-L-(Dox) (SEQ ID NO: 283);
- Ac-G-S-G-R-S-nL-(Dox) (SEQ ID NO: 284);
- Ac-G-T-G-R-S-nL-(Dox) (SEQ ID NO: 285);
- Succ-bA-T-G-R-S-nL-(Dox) (SEQ ID NO: 286);
- Ac-G-T-G-R-S-hCha-(Dox) (SEQ ID NO: 287);
- Ac-G-hS-G-R-S-nL-(Dox) (SEQ ID NO: 288);
- Ac-G-dS-A-R-S-A-(Dox) (SEQ ID NO: 289);
- Ac-G-S-A-R-S-L-(Dox) (SEQ ID NO: 290);
- Ac-G-S-A-R-S-S-Cha-(Dox) (SEQ ID NO: 291);
- Ac-G-S-A-R-S-S-L-(Dox) (SEQ ID NO: 292);
- Ac-G-S-A-R-A-S-L-(Dox) (SEQ ID NO: 293);
- Ac-V-S-G-R-S-L-(Dox) (SEQ ID NO: 294);
- Ac-V-S-G-R-A-L-(Dox) (SEQ ID NO: 295);
- Ac-V-S-G-R-A-S-L-(Dox) (SEQ ID NO: 296);
- Ac-V-S-G-R-S-S-L-(Dox) (SEQ ID NO: 297);
- Ac-V-S-A-R-M-A-(Dox) (SEQ ID NO: 298);
- Ac-V-S-A-R-nL-A-(Dox) (SEQ ID NO: 299);
- Ac-V-S-A-R-S-nL-(Dox) (SEQ ID NO: 300);
- Ac-V-S-A-R-S-L-(Dox) (SEQ ID NO: 301);
- Ac-(Me)C-P-G-R-V-V-(Dox) (SEQ ID NO: 302);
- Ac-(Me)C-P-G-R-V-V-(Dox) (SEQ ID NO: 303);
- Ac-C(Me)-P-G-R-A-L-(Dox) (SEQ ID NO: 304);
- Ac-C(Me)-P-G-R-S-L-(Dox) (SEQ ID NO: 305);
- Ac-C(Me)-P-A-R-S-L-(Dox) (SEQ ID NO: 306);
- Ac-C(Me)-P-A-R-A-S-L-(Dox) (SEQ ID NO: 307);
- Ac-G(tBu)-P-G-R-S-L-(Dox) (SEQ ID NO: 308);
- Ac-Q-S-R-A-A-(taxol) (SEQ ID NO: 552);
- Ac-Q-S-R-S-A-(taxol) (SEQ ID NO: 553);
- Ac-Q-S-R-S-G-(taxol) (SEQ ID NO: 554);
- Ac-R-S-R-A-A-(taxol) (SEQ ID NO: 555);
- Ac-R-Q-S-R-A-A-(taxol) (SEQ ID NO: 556);
- Ac-R-Q-S-R-S-A-(taxol) (SEQ ID NO: 557);
- Ac-R-Q-S-R-S-A-A-(taxol) (SEQ ID NO: 558);
- Ac-R-G-S-G-R-S-A-(taxol) (SEQ ID NO: 559);
- Ac-S-G-R-A-A-(taxol) (SEQ ID NO: 560);
- Ac-S-G-R-S-A-(taxol) (SEQ ID NO: 561);
- Ac-S-G-R-S-S-A-(taxol) (SEQ ID NO: 562);
- Ac-S-G-R-A-S-A-(taxol) (SEQ ID NO: 563);
- Ac-S-G-R-S-G-(taxol) (SEQ ID NO: 564);
- Ac-S-G-R-S-S-G-(taxol) (SEQ ID NO: 565);
- Ac-S-G-R-S-G-A-(taxol) (SEQ ID NO: 566);
- Ac-S-G-R-S-G-G-(taxol) (SEQ ID NO:567);
- Ac-G-T-G-R-S-G-G-(taxol) (SEQ ID NO: 568);
- Ac-L-R-R-Q-S-R-A-A-(Dox) (SEQ ID NO: 597);
- MeSO2-dA(Chx)-Abu-R-S-L-(Dox) (SEQ ID NO: 598);
- Ac-R-A-R-S-L-(Dox) (SEQ ID NO: 599);
- Ac-dA(Chx)-Abu-R-S-L-(Dox) (SEQ ID NO: 600);
- Ac-dA(Chx)-Abu-R-S-S-L-(Dox) (SEQ ID NO: 601);
- Ac-Q-G-R-S-S-L-(Dox) (SEQ ID NO: 602);
- MeOCO-dhF-P(OH)-R-S-S-L-(Dox) (SEQ ID NO: 603);
- MeOCO-Quat4-G-R-S-L-(Dox) (SEQ ID NO: 604);
- As-dCha-P(OH)-R-S-S-L-(Dox) (SEQ ID NO: 605);
- Ac-dCha-Abu-R-S-S-A-(taxol) (SEQ ID NO: 606);
- MeOCO-Quat2-G-R-S-L-NH2 (SEQ ID NO: 607);
- MeOCO-Quat3-G-R-S-L-NH2 (SEQ ID NO: 608); and
- MeOCO-Quat-G-R-S-L-NH2 (SEQ ID NO: 609).
- Naïve or tumor bearing nude mice 8-12 weeks of age have been used for pharmacokinetic studies of the test conjugates. Tumor cells for implantation have been prepared following one of three protocols.
- Protocol A Tumor Cells Collected from Tissue Culture
- Tumor cells are trypsinized and resuspended in the growth medium and centrifuged for 6 min at 200×g. The cells are resuspended in serum-free medium and counted. The appropriate volume of the solution containing the desired number of cells is then transferred to a conical centrifuge tube, centrifuged as before and resuspended in the appropriate volume of a cold 1:1 mixture of cells in phenol free medium:matrigel. Each mouse is inoculated with 0.2-0.5 mL containing between 1×106 and 1×107 tumor cells subcutaneously or orthotopically.
- Protocol B Tumor Cell Suspension Established tumors (200-1000 mm3) are dissected from mice, weighed and rinsed in tumor cell growth medium. The tumors are passed through a steel cell dissociation sieve. The cells are rinsed through the sieve with growth medium. The cells are centrifuged for 6 min at 200×g and resuspended in the appropriate volume of a cold 1:1 mixture of cells:matrigel. Each mouse is inoculated with 0.2-0.5 mL of tumor cells subcutaneously or orthotopically.
- Protocol C Tumor Fragments
- Alternatively a tumor measuring approximately 800 mm3 is dissected out of a mouse, rinsed in tumor cell growth medium and cut into 1-2 mm3 fragments. Each fragment is inoculated subcutaneously or orthotopically using a trocar needle.
- Pharmacokinetic Study
- Naïve or tumor bearing mice are individually weighed and assigned to groups. The mice are dosed with 1-100 umole/kg, including 30 umole/kg, 25 umole/kg, or 21.5 umole/kg of the test conjugate intraperitoneally or intravenously. At a given time point between 5 minutes and 24 hours after administration of the compound the mice are sacrificed. Blood is collected in a syringe containing protease inhibitors such as EDTA, AEBSF, Aprotinin, Leupeptin, Bestatin, Pepstanin A or E64 and transferred into a heparinized blood collection tube. The plasma is prepared by centrifugation. The tumors are collected and pulverized in liquid nitrogen. The resulting tumor powders are stored at −80° C. The tumor powders and plasma are extracted and analyzed for the parent test conjugate and its products including Leucine-doxorubicin (or norleucine-doxorubicin, etc.) and doxorubicin.
- Looking at the delivery of the toxin to the tumor cells, and also looking at the parent conjugate and the levels of toxin (dox and nor-leu dox) in the plasma.
- Results
- For example, test conjugate (21.5 umole/kg of Ac-Gly-Ser-Gly-Arg-Ser-nLeu-Dox (see Example 2)) was administered to naive and tumor bearing (TB) mice intraperitoneally (IP) or intravenously (IV). One hour after administration plasma and tumor tissue was collected from the mice. Concentrations of the test conjugate and its products are compared. The results show that the conjugate does not get into the tumor, the toxins (norleu dox and dox—μM concentrations in the tumor at one hour following the single (both IP and IV) injection. There were lower levels of dox and nor-leu dox the plasma than in the tumor.
- Extraction, Chromatography LC/MS Conditions
- Plasma: Plasma samples are prepared using acetonitrile protein precipitation. A standard curve was constructed from addition of 5 to 20 μL volumes of a standard compound to 0.1 mL or 0.05 mL volumes of plasma on ice. The standard curve ranges from 10 ng/mL-1 ug/mL or from 100 ng/mL-4 ug/mL of the standard compound. Immediumtely after standard addition, acetonitrile is added to precipitate the proteins. The study plasma samples were prepared by thawing the frozen plasma samples on ice. The aliquots were added directly to the acetonitrile. After sample precipitation, the sample is mixed using vortex mixing. The precipitate was pelleted using centrifugation. The supernatent was dried using vacuum centrifugation. The sample was reconstituted with 0.15 mL of 30% acetonitrile—70% (0.01 M ammonium acetate with 0.1% formic acid). 0.01 mL of the sample was injected for LC-MS analysis. The HPLC conditions were a linear gradient of 20% acetonitrile—80% (10 mM ammonium acetate—0.1% formic acid) to 50% acetonitrile—50% (10 mM ammonium acetate—0.1% formic acid) in 1 minute at 0.3 mL/min in a 30×2.1 mm Zorbax SB C18 HPLC column. Detection was provided by a triple quad mass spectrometer with electrospray ionization. Doxorubicin was monitored using the m/z transition 544.1-396.8. Leucine-doxorubicin was monitored using 657.2-242.8. An exemplary parent conjugate was monitored using 1555.9-1555.9. Scanning LC-MS and fluorescence detection was used to identify cleavage products other than doxorubicin or leucine-doxorubicin (or norleucine-doxorubicin, etc.) in the plasma.
- Tumor: Immediately after excision from the mouse, the tumor for analysis is weighed and placed into a mortar containing liquid nitrogen. With the mortar nested in a bed of dry ice, the tumor is ground into a fine powder while additional liquid nitrogen is added as needed to avoid thawing. When a homogeneous tumor powder is achieved, the remaining liquid nitrogen is allowed to boil off. The tumor powder is quantitatively transferred to a 15 ml conical tube that has been pre-chilled and is on dry ice. The sample is stored at −70° C. until analysis. The tumor powder is thawed on ice and vortex mixed with 0.01 M ammonium acetate in a 1 gram tumor/mL ammonium acetate solution concentration to form a slurry. An aliquot of 0.1 mL of the tumor slurry is precipitated with 0.5 mL acetonitrile. The supernatant is separated from the precipitated solids and then evaporated using vacuum centrifugation. Quantification of doxorubicin, leucine-doxorubicin (or norleucine-docorubicin, etc.), is achieved by reference to a standard curve constructed from spiking measured amounts of standard compounds (doxorubicin, leucine-doxorubicin, etc.) into control tumor slurry. A typical standard curve ranges from 1 ng to 200 ng of compound per aliquot of tumor slurry. After the unknown samples and standards are processed and dried, the residue is reconstituted in 0.15 mL of 30% acetonitrile—70% (0.01 M ammonium acetate+0.1% formic acid). 10 μL of solution is injected onto a liquid chromatography—mass spectrometry system. The HPLC conditions were a linear gradient of 20% acetonitrile—80% (10 mM ammonium acetate—0.1% formic acid) to 50% acetonitrile—50% (10 mM ammonium acetate—0.1% formic acid) in 1 minute at 0.3 mL/min in a 30×2.1 mm Zorbax SB C18 HPLC column. Detection was provided by a triple quad mass spectrometer with electrospray ionization. Doxorubicin was monitored using the m/z transition 544.1-396.8. Leucine-doxorubicin was monitored using 657.2-242.8.
- Since modifications will be apparent to those of skill in this art, it is intended that this invention be limited only by the scope of the appended claims.
-
0 SEQUENCE LISTING <160> NUMBER OF SEQ ID NOS: 611 <210> SEQ ID NO 1 <211> LENGTH: 3147 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (23)...(2589) <223> OTHER INFORMATION: Nucleotide sequence encoding MTSP1 <300> PUBLICATION INFORMATION: <301> AUTHORS: O′Brien, T.J. and Tanimoto, H. <308> DATABASE ACCESSION NUMBER: GenBank AR081724 <309> DATABASE ENTRY DATE: 2000-08-31 <310> PATENT DOCUMENT NUMBER: US Pat 5972616 <311> PATENT FILING DATE: 1998-02-20 <312> PUBLICATION DATE: 1999-10-26 <400> SEQUENCE: 1 tcaagagcgg cctcggggta cc atg ggg agc gat cgg gcc cgc aag ggc gga 52 Met Gly Ser Asp Arg Ala Arg Lys Gly Gly 1 5 10 ggg ggc ccg aag gac ttc ggc gcg gga ctc aag tac aac tcc cgg cac 100 Gly Gly Pro Lys Asp Phe Gly Ala Gly Leu Lys Tyr Asn Ser Arg His 15 20 25 gag aaa gtg aat ggc ttg gag gaa ggc gtg gag ttc ctg cca gtc aac 148 Glu Lys Val Asn Gly Leu Glu Glu Gly Val Glu Phe Leu Pro Val Asn 30 35 40 aac gtc aag aag gtg gaa aag cat ggc ccg ggg cgc tgg gtg gtg ctg 196 Asn Val Lys Lys Val Glu Lys His Gly Pro Gly Arg Trp Val Val Leu 45 50 55 gca gcc gtg ctg atc ggc ctc ctc ttg gtc ttg ctg ggg atc ggc ttc 244 Ala Ala Val Leu Ile Gly Leu Leu Leu Val Leu Leu Gly Ile Gly Phe 60 65 70 ctg gtg tgg cat ttg cag tac cgg gac gtg cgt gtc cag aag gtc ttc 292 Leu Val Trp His Leu Gln Tyr Arg Asp Val Arg Val Gln Lys Val Phe 75 80 85 90 aat ggc tac atg agg atc aca aat gag aat ttt gtg gat gcc tac gag 340 Asn Gly Tyr Met Arg Ile Thr Asn Glu Asn Phe Val Asp Ala Tyr Glu 95 100 105 aac tcc aac tcc act gag ttt gta agc ctg gcc agc aag gtg aag gac 388 Asn Ser Asn Ser Thr Glu Phe Val Ser Leu Ala Ser Lys Val Lys Asp 110 115 120 gcg ctg aag ctg ctg tac agc gga gtc cca ttc ctg ggc ccc tac cac 436 Ala Leu Lys Leu Leu Tyr Ser Gly Val Pro Phe Leu Gly Pro Tyr His 125 130 135 aag gag tcg gct gtg acg gcc ttc agc gag ggc agc gtc atc gcc tac 484 Lys Glu Ser Ala Val Thr Ala Phe Ser Glu Gly Ser Val Ile Ala Tyr 140 145 150 tac tgg tct gag ttc agc atc ccg cag cac ctg gtg gag gag gcc gag 532 Tyr Trp Ser Glu Phe Ser Ile Pro Gln His Leu Val Glu Glu Ala Glu 155 160 165 170 cgc gtc atg gcc gag gag cgc gta gtc atg ctg ccc ccg cgg gcg cgc 580 Arg Val Met Ala Glu Glu Arg Val Val Met Leu Pro Pro Arg Ala Arg 175 180 185 tcc ctg aag tcc ttt gtg gtc acc tca gtg gtg gct ttc ccc acg gac 628 Ser Leu Lys Ser Phe Val Val Thr Ser Val Val Ala Phe Pro Thr Asp 190 195 200 tcc aaa aca gta cag agg acc cag gac aac agc tgc agc ttt ggc ctg 676 Ser Lys Thr Val Gln Arg Thr Gln Asp Asn Ser Cys Ser Phe Gly Leu 205 210 215 cac gcc cgc ggt gtg gag ctg atg cgc ttc acc acg ccc ggc ttc cct 724 His Ala Arg Gly Val Glu Leu Met Arg Phe Thr Thr Pro Gly Phe Pro 220 225 230 gac agc ccc tac ccc gct cat gcc cgc tgc cag tgg gcc ctg cgg ggg 772 Asp Ser Pro Tyr Pro Ala His Ala Arg Cys Gln Trp Ala Leu Arg Gly 235 240 245 250 gac gcc gac tca gtg ctg agc ctc acc ttc cgc agc ttt gac ctt gcg 820 Asp Ala Asp Ser Val Leu Ser Leu Thr Phe Arg Ser Phe Asp Leu Ala 255 260 265 tcc tgc gac gag cgc ggc agc gac ctg gtg acg gtg tac aac acc ctg 868 Ser Cys Asp Glu Arg Gly Ser Asp Leu Val Thr Val Tyr Asn Thr Leu 270 275 280 agc ccc atg gag ccc cac gcc ctg gtg cag ttg tgt ggc acc tac cct 916 Ser Pro Met Glu Pro His Ala Leu Val Gln Leu Cys Gly Thr Tyr Pro 285 290 295 ccc tcc tac aac ctg acc ttc cac tcc tcc cag aac gtc ctg ctc atc 964 Pro Ser Tyr Asn Leu Thr Phe His Ser Ser Gln Asn Val Leu Leu Ile 300 305 310 aca ctg ata acc aac act gag cgg cgg cat ccc ggc ttt gag gcc acc 1012 Thr Leu Ile Thr Asn Thr Glu Arg Arg His Pro Gly Phe Glu Ala Thr 315 320 325 330 ttc ttc cag ctg cct agg atg agc agc tgt gga ggc cgc tta cgt aaa 1060 Phe Phe Gln Leu Pro Arg Met Ser Ser Cys Gly Gly Arg Leu Arg Lys 335 340 345 gcc cag ggg aca ttc aac agc ccc tac tac cca ggc cac tac cca ccc 1108 Ala Gln Gly Thr Phe Asn Ser Pro Tyr Tyr Pro Gly His Tyr Pro Pro 350 355 360 aac att gac tgc aca tgg aac att gag gtg ccc aac aac cag cat gtg 1156 Asn Ile Asp Cys Thr Trp Asn Ile Glu Val Pro Asn Asn Gln His Val 365 370 375 aag gtg agc ttc aaa ttc ttc tac ctg ctg gag ccc ggc gtg cct gcg 1204 Lys Val Ser Phe Lys Phe Phe Tyr Leu Leu Glu Pro Gly Val Pro Ala 380 385 390 ggc acc tgc ccc aag gac tac gtg gag atc aat ggg gag aaa tac tgc 1252 Gly Thr Cys Pro Lys Asp Tyr Val Glu Ile Asn Gly Glu Lys Tyr Cys 395 400 405 410 gga gag agg tcc cag ttc gtc gtc acc agc aac agc aac aag atc aca 1300 Gly Glu Arg Ser Gln Phe Val Val Thr Ser Asn Ser Asn Lys Ile Thr 415 420 425 gtt cgc ttc cac tca gat cag tcc tac acc gac acc ggc ttc tta gct 1348 Val Arg Phe His Ser Asp Gln Ser Tyr Thr Asp Thr Gly Phe Leu Ala 430 435 440 gaa tac ctc tcc tac gac tcc agt gac cca tgc ccg ggg cag ttc acg 1396 Glu Tyr Leu Ser Tyr Asp Ser Ser Asp Pro Cys Pro Gly Gln Phe Thr 445 450 455 tgc cgc acg ggg cgg tgt atc cgg aag gag ctg cgc tgt gat ggc tgg 1444 Cys Arg Thr Gly Arg Cys Ile Arg Lys Glu Leu Arg Cys Asp Gly Trp 460 465 470 gcc gac tgc acc gac cac agc gat gag ctc aac tgc agt tgc gac gcc 1492 Ala Asp Cys Thr Asp His Ser Asp Glu Leu Asn Cys Ser Cys Asp Ala 475 480 485 490 ggc cac cag ttc acg tgc aag aac aag ttc tgc aag ccc ctc ttc tgg 1540 Gly His Gln Phe Thr Cys Lys Asn Lys Phe Cys Lys Pro Leu Phe Trp 495 500 505 gtc tgc gac agt gtg aac gac tgc gga gac aac agc gac gag cag ggg 1588 Val Cys Asp Ser Val Asn Asp Cys Gly Asp Asn Ser Asp Glu Gln Gly 510 515 520 tgc agt tgt ccg gcc cag acc ttc agg tgt tcc aat ggg aag tgc ctc 1636 Cys Ser Cys Pro Ala Gln Thr Phe Arg Cys Ser Asn Gly Lys Cys Leu 525 530 535 tcg aaa agc cag cag tgc aat ggg aag gac gac tgt ggg gac ggg tcc 1684 Ser Lys Ser Gln Gln Cys Asn Gly Lys Asp Asp Cys Gly Asp Gly Ser 540 545 550 gac gag gcc tcc tgc ccc aag gtg aac gtc gtc act tgt acc aaa cac 1732 Asp Glu Ala Ser Cys Pro Lys Val Asn Val Val Thr Cys Thr Lys His 555 560 565 570 acc tac cgc tgc ctc aat ggg ctc tgc ttg agc aag ggc aac cct gag 1780 Thr Tyr Arg Cys Leu Asn Gly Leu Cys Leu Ser Lys Gly Asn Pro Glu 575 580 585 tgt gac ggg aag gag gac tgt agc gac ggc tca gat gag aag gac tgc 1828 Cys Asp Gly Lys Glu Asp Cys Ser Asp Gly Ser Asp Glu Lys Asp Cys 590 595 600 gac tgt ggg ctg cgg tca ttc acg aga cag gct cgt gtt gtt ggg ggc 1876 Asp Cys Gly Leu Arg Ser Phe Thr Arg Gln Ala Arg Val Val Gly Gly 605 610 615 acg gat gcg gat gag ggc gag tgg ccc tgg cag gta agc ctg cat gct 1924 Thr Asp Ala Asp Glu Gly Glu Trp Pro Trp Gln Val Ser Leu His Ala 620 625 630 ctg ggc cag ggc cac atc tgc ggt gct tcc ctc atc tct ccc aac tgg 1972 Leu Gly Gln Gly His Ile Cys Gly Ala Ser Leu Ile Ser Pro Asn Trp 635 640 645 650 ctg gtc tct gcc gca cac tgc tac atc gat gac aga gga ttc agg tac 2020 Leu Val Ser Ala Ala His Cys Tyr Ile Asp Asp Arg Gly Phe Arg Tyr 655 660 665 tca gac ccc acg cag tgg acg gcc ttc ctg ggc ttg cac gac cag agc 2068 Ser Asp Pro Thr Gln Trp Thr Ala Phe Leu Gly Leu His Asp Gln Ser 670 675 680 cag cgc agc gcc cct ggg gtg cag gag cgc agg ctc aag cgc atc atc 2116 Gln Arg Ser Ala Pro Gly Val Gln Glu Arg Arg Leu Lys Arg Ile Ile 685 690 695 tcc cac ccc ttc ttc aat gac ttc acc ttc gac tat gac atc gcg ctg 2164 Ser His Pro Phe Phe Asn Asp Phe Thr Phe Asp Tyr Asp Ile Ala Leu 700 705 710 ctg gag ctg gag aaa ccg gca gag tac agc tcc atg gtg cgg ccc atc 2212 Leu Glu Leu Glu Lys Pro Ala Glu Tyr Ser Ser Met Val Arg Pro Ile 715 720 725 730 tgc ctg ccg gac gcc tcc cat gtc ttc cct gcc ggc aag gcc atc tgg 2260 Cys Leu Pro Asp Ala Ser His Val Phe Pro Ala Gly Lys Ala Ile Trp 735 740 745 gtc acg ggc tgg gga cac acc cag tat gga ggc act ggc gcg ctg atc 2308 Val Thr Gly Trp Gly His Thr Gln Tyr Gly Gly Thr Gly Ala Leu Ile 750 755 760 ctg caa aag ggt gag atc cgc gtc atc aac cag acc acc tgc gag aac 2356 Leu Gln Lys Gly Glu Ile Arg Val Ile Asn Gln Thr Thr Cys Glu Asn 765 770 775 ctc ctg ccg cag cag atc acg ccg cgc atg atg tgc gtg ggc ttc ctc 2404 Leu Leu Pro Gln Gln Ile Thr Pro Arg Met Met Cys Val Gly Phe Leu 780 785 790 agc ggc ggc gtg gac tcc tgc cag ggt gat tcc ggg gga ccc ctg tcc 2452 Ser Gly Gly Val Asp Ser Cys Gln Gly Asp Ser Gly Gly Pro Leu Ser 795 800 805 810 agc gtg gag gcg gat ggg cgg atc ttc cag gcc ggt gtg gtg agc tgg 2500 Ser Val Glu Ala Asp Gly Arg Ile Phe Gln Ala Gly Val Val Ser Trp 815 820 825 gga gac ggc tgc gct cag agg aac aag cca ggc gtg tac aca agg ctc 2548 Gly Asp Gly Cys Ala Gln Arg Asn Lys Pro Gly Val Tyr Thr Arg Leu 830 835 840 cct ctg ttt cgg gac tgg atc aaa gag aac act ggg gta ta ggggccgggg 2599 Pro Leu Phe Arg Asp Trp Ile Lys Glu Asn Thr Gly Val 845 850 855 ccacccaaat gtgtacacct gcggggccac ccatcgtcca ccccagtgtg cacgcctgca 2659 ggctggagac tggaccgctg actgcaccag cgcccccaga acatacactg tgaactcaat 2719 ctccagggct ccaaatctgc ctagaaaacc tctcgcttcc tcagcctcca aagtggagct 2779 gggaggtaga aggggaggac actggtggtt ctactgaccc aactgggggc aaaggtttga 2839 agacacagcc tcccccgcca gccccaagct gggccgaggc gcgtttgtgt atatctgcct 2899 cccctgtctg taaggagcag cgggaacgga gcttcggagc ctcctcagtg aaggtggtgg 2959 ggctgccgga tctgggctgt ggggcccttg ggccacgctc ttgaggaagc ccaggctcgg 3019 aggaccctgg aaaacagacg ggtctgagac tgaaattgtt ttaccagctc ccagggtgga 3079 cttcagtgtg tgtatttgtg taaatgggta aaacaattta tttcttttta aaaaaaaaaa 3139 aaaaaaaa 3147 <210> SEQ ID NO 2<211> LENGTH: 855 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 2 Met Gly Ser Asp Arg Ala Arg Lys Gly Gly Gly Gly Pro Lys Asp Phe 1 5 10 15 Gly Ala Gly Leu Lys Tyr Asn Ser Arg His Glu Lys Val Asn Gly Leu 20 25 30 Glu Glu Gly Val Glu Phe Leu Pro Val Asn Asn Val Lys Lys Val Glu 35 40 45 Lys His Gly Pro Gly Arg Trp Val Val Leu Ala Ala Val Leu Ile Gly 50 55 60 Leu Leu Leu Val Leu Leu Gly Ile Gly Phe Leu Val Trp His Leu Gln 65 70 75 80 Tyr Arg Asp Val Arg Val Gln Lys Val Phe Asn Gly Tyr Met Arg Ile 85 90 95 Thr Asn Glu Asn Phe Val Asp Ala Tyr Glu Asn Ser Asn Ser Thr Glu 100 105 110 Phe Val Ser Leu Ala Ser Lys Val Lys Asp Ala Leu Lys Leu Leu Tyr 115 120 125 Ser Gly Val Pro Phe Leu Gly Pro Tyr His Lys Glu Ser Ala Val Thr 130 135 140 Ala Phe Ser Glu Gly Ser Val Ile Ala Tyr Tyr Trp Ser Glu Phe Ser 145 150 155 160 Ile Pro Gln His Leu Val Glu Glu Ala Glu Arg Val Met Ala Glu Glu 165 170 175 Arg Val Val Met Leu Pro Pro Arg Ala Arg Ser Leu Lys Ser Phe Val 180 185 190 Val Thr Ser Val Val Ala Phe Pro Thr Asp Ser Lys Thr Val Gln Arg 195 200 205 Thr Gln Asp Asn Ser Cys Ser Phe Gly Leu His Ala Arg Gly Val Glu 210 215 220 Leu Met Arg Phe Thr Thr Pro Gly Phe Pro Asp Ser Pro Tyr Pro Ala 225 230 235 240 His Ala Arg Cys Gln Trp Ala Leu Arg Gly Asp Ala Asp Ser Val Leu 245 250 255 Ser Leu Thr Phe Arg Ser Phe Asp Leu Ala Ser Cys Asp Glu Arg Gly 260 265 270 Ser Asp Leu Val Thr Val Tyr Asn Thr Leu Ser Pro Met Glu Pro His 275 280 285 Ala Leu Val Gln Leu Cys Gly Thr Tyr Pro Pro Ser Tyr Asn Leu Thr 290 295 300 Phe His Ser Ser Gln Asn Val Leu Leu Ile Thr Leu Ile Thr Asn Thr 305 310 315 320 Glu Arg Arg His Pro Gly Phe Glu Ala Thr Phe Phe Gln Leu Pro Arg 325 330 335 Met Ser Ser Cys Gly Gly Arg Leu Arg Lys Ala Gln Gly Thr Phe Asn 340 345 350 Ser Pro Tyr Tyr Pro Gly His Tyr Pro Pro Asn Ile Asp Cys Thr Trp 355 360 365 Asn Ile Glu Val Pro Asn Asn Gln His Val Lys Val Ser Phe Lys Phe 370 375 380 Phe Tyr Leu Leu Glu Pro Gly Val Pro Ala Gly Thr Cys Pro Lys Asp 385 390 395 400 Tyr Val Glu Ile Asn Gly Glu Lys Tyr Cys Gly Glu Arg Ser Gln Phe 405 410 415 Val Val Thr Ser Asn Ser Asn Lys Ile Thr Val Arg Phe His Ser Asp 420 425 430 Gln Ser Tyr Thr Asp Thr Gly Phe Leu Ala Glu Tyr Leu Ser Tyr Asp 435 440 445 Ser Ser Asp Pro Cys Pro Gly Gln Phe Thr Cys Arg Thr Gly Arg Cys 450 455 460 Ile Arg Lys Glu Leu Arg Cys Asp Gly Trp Ala Asp Cys Thr Asp His 465 470 475 480 Ser Asp Glu Leu Asn Cys Ser Cys Asp Ala Gly His Gln Phe Thr Cys 485 490 495 Lys Asn Lys Phe Cys Lys Pro Leu Phe Trp Val Cys Asp Ser Val Asn 500 505 510 Asp Cys Gly Asp Asn Ser Asp Glu Gln Gly Cys Ser Cys Pro Ala Gln 515 520 525 Thr Phe Arg Cys Ser Asn Gly Lys Cys Leu Ser Lys Ser Gln Gln Cys 530 535 540 Asn Gly Lys Asp Asp Cys Gly Asp Gly Ser Asp Glu Ala Ser Cys Pro 545 550 555 560 Lys Val Asn Val Val Thr Cys Thr Lys His Thr Tyr Arg Cys Leu Asn 565 570 575 Gly Leu Cys Leu Ser Lys Gly Asn Pro Glu Cys Asp Gly Lys Glu Asp 580 585 590 Cys Ser Asp Gly Ser Asp Glu Lys Asp Cys Asp Cys Gly Leu Arg Ser 595 600 605 Phe Thr Arg Gln Ala Arg Val Val Gly Gly Thr Asp Ala Asp Glu Gly 610 615 620 Glu Trp Pro Trp Gln Val Ser Leu His Ala Leu Gly Gln Gly His Ile 625 630 635 640 Cys Gly Ala Ser Leu Ile Ser Pro Asn Trp Leu Val Ser Ala Ala His 645 650 655 Cys Tyr Ile Asp Asp Arg Gly Phe Arg Tyr Ser Asp Pro Thr Gln Trp 660 665 670 Thr Ala Phe Leu Gly Leu His Asp Gln Ser Gln Arg Ser Ala Pro Gly 675 680 685 Val Gln Glu Arg Arg Leu Lys Arg Ile Ile Ser His Pro Phe Phe Asn 690 695 700 Asp Phe Thr Phe Asp Tyr Asp Ile Ala Leu Leu Glu Leu Glu Lys Pro 705 710 715 720 Ala Glu Tyr Ser Ser Met Val Arg Pro Ile Cys Leu Pro Asp Ala Ser 725 730 735 His Val Phe Pro Ala Gly Lys Ala Ile Trp Val Thr Gly Trp Gly His 740 745 750 Thr Gln Tyr Gly Gly Thr Gly Ala Leu Ile Leu Gln Lys Gly Glu Ile 755 760 765 Arg Val Ile Asn Gln Thr Thr Cys Glu Asn Leu Leu Pro Gln Gln Ile 770 775 780 Thr Pro Arg Met Met Cys Val Gly Phe Leu Ser Gly Gly Val Asp Ser 785 790 795 800 Cys Gln Gly Asp Ser Gly Gly Pro Leu Ser Ser Val Glu Ala Asp Gly 805 810 815 Arg Ile Phe Gln Ala Gly Val Val Ser Trp Gly Asp Gly Cys Ala Gln 820 825 830 Arg Asn Lys Pro Gly Val Tyr Thr Arg Leu Pro Leu Phe Arg Asp Trp 835 840 845 Ile Lys Glu Asn Thr Gly Val 850 855 <210> SEQ ID NO 3<211> LENGTH: 2137 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (261)...(1574) <223> OTHER INFORMATION: Nucleic acid encoding a transmembrane serine protease (MTSP3) protein <400> SEQUENCE: 3 ccatcctaat acgactcact atagggctcg agcggccgcc cgggcaggtc agagagaggc 60 agcagcttgc tcagcggaca aggatgctgg gcgtgaggga ccaaggcctg ccctgcactc 120 gggcctcctc cagccagtgc tgaccaggga cttctgacct gctggccagc caggacctgt 180 gtggggaggc cctcctgctg ccttggggtg acaatctcag ctccaggcta cagggagacc 240 gggaggatca cagagccagc atg tta cag gat cct gac agt gat caa cct ctg 293Met Leu Gln Asp Pro Asp Ser Asp Gln Pro Leu 1 5 10 aac agc ctc gat gtc aaa ccc ctg cgc aaa ccc cgt atc ccc atg gag 341 Asn Ser Leu Asp Val Lys Pro Leu Arg Lys Pro Arg Ile Pro Met Glu 15 20 25 acc ttc aga aag gtg ggg atc ccc atc atc ata gca cta ctg agc ctg 389Thr Phe Arg Lys Val Gly Ile Pro Ile Ile Ile Ala Leu Leu Ser Leu 30 35 40 gcg agt atc atc att gtg gtt gtc ctc atc aag gtg att ctg gat aaa 437 Ala Ser Ile Ile Ile Val Val Val Leu Ile Lys Val Ile Leu Asp Lys 45 50 55 tac tac ttc ctc tgc ggg cag cct ctc cac ttc atc ccg agg aag cag 485Tyr Tyr Phe Leu Cys Gly Gln Pro Leu His Phe Ile Pro Arg Lys Gln 60 65 70 75 ctg tgt gac gga gag ctg gac tgt ccc ttg ggg gag gac gag gag cac 533Leu Cys Asp Gly Glu Leu Asp Cys Pro Leu Gly Glu Asp Glu Glu His 80 85 90 tgt gtc aag agc ttc ccc gaa ggg cct gca gtg gca gtc cgc ctc tcc 581 Cys Val Lys Ser Phe Pro Glu Gly Pro Ala Val Ala Val Arg Leu Ser 95 100 105 aag gac cga tcc aca ctg cag gtg ctg gac tcg gcc aca ggg aac tgg 629 Lys Asp Arg Ser Thr Leu Gln Val Leu Asp Ser Ala Thr Gly Asn Trp 110 115 120 ttc tct gcc tgt ttc gac aac ttc aca gaa gct ctc gct gag aca gcc 677 Phe Ser Ala Cys Phe Asp Asn Phe Thr Glu Ala Leu Ala Glu Thr Ala 125 130 135 tgt agg cag atg ggc tac agc agc aaa ccc acc ttc aga gct gtg gag 725 Cys Arg Gln Met Gly Tyr Ser Ser Lys Pro Thr Phe Arg Ala Val Glu 140 145 150 155 att ggc cca gac cag gat ctg gat gtt gtt gaa atc aca gaa aac agc 773 Ile Gly Pro Asp Gln Asp Leu Asp Val Val Glu Ile Thr Glu Asn Ser 160 165 170 cag gag ctt cgc atg cgg aac tca agt ggg ccc tgt ctc tca ggc tcc 821 Gln Glu Leu Arg Met Arg Asn Ser Ser Gly Pro Cys Leu Ser Gly Ser 175 180 185 ctg gtc tcc ctg cac tgt ctt gcc tgt ggg aag agc ctg aag acc ccc 869 Leu Val Ser Leu His Cys Leu Ala Cys Gly Lys Ser Leu Lys Thr Pro 190 195 200 cgt gtg gtg ggt ggg gag gag gcc tct gtg gat tct tgg cct tgg cag 917 Arg Val Val Gly Gly Glu Glu Ala Ser Val Asp Ser Trp Pro Trp Gln 205 210 215 gtc agc atc cag tac gac ata cag cac gtc tgt gga ggg agc atc ctg 965 Val Ser Ile Gln Tyr Asp Ile Gln His Val Cys Gly Gly Ser Ile Leu 220 225 230 235 gac ccc cac tgg gtc ctc acg gca gcc cac tgc ttc agg aaa cat acc 1013 Asp Pro His Trp Val Leu Thr Ala Ala His Cys Phe Arg Lys His Thr 240 245 250 gat gtg ttc aac tgg aag gtg cgg gca ggc tca gac aaa ctg ggc agc 1061 Asp Val Phe Asn Trp Lys Val Arg Ala Gly Ser Asp Lys Leu Gly Ser 255 260 265 ttc cca tcc ctg gct gtg gcc aag atc atc atc att gaa ttc aac ccc 1109 Phe Pro Ser Leu Ala Val Ala Lys Ile Ile Ile Ile Glu Phe Asn Pro 270 275 280 atg tac ccc aaa gac aat gac atc gcc ctc atg aag ctg cag ttc cca 1157 Met Tyr Pro Lys Asp Asn Asp Ile Ala Leu Met Lys Leu Gln Phe Pro 285 290 295 ctc act ttc tca ggc aca gtc agg ctc atc tgt ctg ccc ttc ttt gat 1205 Leu Thr Phe Ser Gly Thr Val Arg Leu Ile Cys Leu Pro Phe Phe Asp 300 305 310 315 gag gag ctc act cca gcc acc cca ctc tgg atc att gga tgg ggc ttt 1253 Glu Glu Leu Thr Pro Ala Thr Pro Leu Trp Ile Ile Gly Trp Gly Phe 320 325 330 acg aag cag aat gga ggg aag atg tct gac ata ctg ctg cag gcg tca 1301 Thr Lys Gln Asn Gly Gly Lys Met Ser Asp Ile Leu Leu Gln Ala Ser 335 340 345 gtc cag gtc att gac agc aca cgg tgc aat gca gac gat gcg tac cag 1349 Val Gln Val Ile Asp Ser Thr Arg Cys Asn Ala Asp Asp Ala Tyr Gln 350 355 360 ggg gaa gtc acc gag aag atg atg tgt gca ggc atc ccg gaa ggg ggt 1397 Gly Glu Val Thr Glu Lys Met Met Cys Ala Gly Ile Pro Glu Gly Gly 365 370 375 gtg gac acc tgc cag ggt gac agt ggt ggg ccc ctg atg tac caa tct 1445 Val Asp Thr Cys Gln Gly Asp Ser Gly Gly Pro Leu Met Tyr Gln Ser 380 385 390 395 gac cag tgg cat gtg gtg ggc atc gtt agc tgg ggc tat ggc tgc ggg 1493 Asp Gln Trp His Val Val Gly Ile Val Ser Trp Gly Tyr Gly Cys Gly 400 405 410 ggc ccg agc acc cca gga gta tac acc aag gtc tca gcc tat ctc aac 1541 Gly Pro Ser Thr Pro Gly Val Tyr Thr Lys Val Ser Ala Tyr Leu Asn 415 420 425 tgg atc tac aat gtc tgg aag gct gag ctg taa tgctgctgcc cctttgcagt 1594 Trp Ile Tyr Asn Val Trp Lys Ala Glu Leu * 430 435 gctgggagcc gcttccttcc tgccctgccc acctggggat cccccaaagt cagacacaga 1654 gcaagagtcc ccttgggtac acccctctgc ccacagcctc agcatttctt ggagcagcaa 1714 agggcctcaa ttcctgtaag agaccctcgc agcccagagg cgcccagagg aagtcagcag 1774 ccctagctcg gccacacttg gtgctcccag catcccaggg agagacacag cccactgaac 1834 aaggtctcag gggtattgct aagccaagaa ggaactttcc cacactactg aatggaagca 1894 ggctgtcttg taaaagccca gatcactgtg ggctggagag gagaaggaaa gggtctgcgc 1954 cagccctgtc cgtcttcacc catccccaag cctactagag caagaaacca gttgtaatat 2014 aaaatgcact gccctactgt tggtatgact accgttacct actgttgtca ttgttattac 2074 agctatggcc actattatta aagagctgtg taacaaaaaa aaaaaaaaaa aaaaaaaaaa 2134 aaa 2137 <210> SEQ ID NO 4<211> LENGTH: 437 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 4 Met Leu Gln Asp Pro Asp Ser Asp Gln Pro Leu Asn Ser Leu Asp Val 1 5 10 15 Lys Pro Leu Arg Lys Pro Arg Ile Pro Met Glu Thr Phe Arg Lys Val 20 25 30 Gly Ile Pro Ile Ile Ile Ala Leu Leu Ser Leu Ala Ser Ile Ile Ile 35 40 45 Val Val Val Leu Ile Lys Val Ile Leu Asp Lys Tyr Tyr Phe Leu Cys 50 55 60 Gly Gln Pro Leu His Phe Ile Pro Arg Lys Gln Leu Cys Asp Gly Glu 65 70 75 80 Leu Asp Cys Pro Leu Gly Glu Asp Glu Glu His Cys Val Lys Ser Phe 85 90 95 Pro Glu Gly Pro Ala Val Ala Val Arg Leu Ser Lys Asp Arg Ser Thr 100 105 110 Leu Gln Val Leu Asp Ser Ala Thr Gly Asn Trp Phe Ser Ala Cys Phe 115 120 125 Asp Asn Phe Thr Glu Ala Leu Ala Glu Thr Ala Cys Arg Gln Met Gly 130 135 140 Tyr Ser Ser Lys Pro Thr Phe Arg Ala Val Glu Ile Gly Pro Asp Gln 145 150 155 160 Asp Leu Asp Val Val Glu Ile Thr Glu Asn Ser Gln Glu Leu Arg Met 165 170 175 Arg Asn Ser Ser Gly Pro Cys Leu Ser Gly Ser Leu Val Ser Leu His 180 185 190 Cys Leu Ala Cys Gly Lys Ser Leu Lys Thr Pro Arg Val Val Gly Gly 195 200 205 Glu Glu Ala Ser Val Asp Ser Trp Pro Trp Gln Val Ser Ile Gln Tyr 210 215 220 Asp Ile Gln His Val Cys Gly Gly Ser Ile Leu Asp Pro His Trp Val 225 230 235 240 Leu Thr Ala Ala His Cys Phe Arg Lys His Thr Asp Val Phe Asn Trp 245 250 255 Lys Val Arg Ala Gly Ser Asp Lys Leu Gly Ser Phe Pro Ser Leu Ala 260 265 270 Val Ala Lys Ile Ile Ile Ile Glu Phe Asn Pro Met Tyr Pro Lys Asp 275 280 285 Asn Asp Ile Ala Leu Met Lys Leu Gln Phe Pro Leu Thr Phe Ser Gly 290 295 300 Thr Val Arg Leu Ile Cys Leu Pro Phe Phe Asp Glu Glu Leu Thr Pro 305 310 315 320 Ala Thr Pro Leu Trp Ile Ile Gly Trp Gly Phe Thr Lys Gln Asn Gly 325 330 335 Gly Lys Met Ser Asp Ile Leu Leu Gln Ala Ser Val Gln Val Ile Asp 340 345 350 Ser Thr Arg Cys Asn Ala Asp Asp Ala Tyr Gln Gly Glu Val Thr Glu 355 360 365 Lys Met Met Cys Ala Gly Ile Pro Glu Gly Gly Val Asp Thr Cys Gln 370 375 380 Gly Asp Ser Gly Gly Pro Leu Met Tyr Gln Ser Asp Gln Trp His Val 385 390 395 400 Val Gly Ile Val Ser Trp Gly Tyr Gly Cys Gly Gly Pro Ser Thr Pro 405 410 415 Gly Val Tyr Thr Lys Val Ser Ala Tyr Leu Asn Trp Ile Tyr Asn Val 420 425 430 Trp Lys Ala Glu Leu 435 <210> SEQ ID NO 5 <211> LENGTH: 708 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (1)...(708) <223> OTHER INFORMATION: Nucleic acid encoding an MTSP4 protease domain <400> SEQUENCE: 5 att gtt ggt gga gct gtg tcc tcc gag ggt gag tgg cca tgg cag gcc 48 Ile Val Gly Gly Ala Val Ser Ser Glu Gly Glu Trp Pro Trp Gln Ala 1 5 10 15 agc ctc cag gtt cgg ggt cga cac atc tgt ggg ggg gcc ctc atc gct 96 Ser Leu Gln Val Arg Gly Arg His Ile Cys Gly Gly Ala Leu Ile Ala 20 25 30 gac cgc tgg gtg ata aca gct gcc cac tgc ttc cag gag gac agc atg 144 Asp Arg Trp Val Ile Thr Ala Ala His Cys Phe Gln Glu Asp Ser Met 35 40 45 gcc tcc acg gtg ctg tgg acc gtg ttc ctg ggc aag gtg tgg cag aac 192 Ala Ser Thr Val Leu Trp Thr Val Phe Leu Gly Lys Val Trp Gln Asn 50 55 60 tcg cgc tgg cct gga gag gtg tcc ttc aag gtg agc cgc ctg ctc ctg 240 Ser Arg Trp Pro Gly Glu Val Ser Phe Lys Val Ser Arg Leu Leu Leu 65 70 75 80 cac ccg tac cac gaa gag gac agc cat gac tac gac gtg gcg ctg ctg 288His Pro Tyr His Glu Glu Asp Ser His Asp Tyr Asp Val Ala Leu Leu 85 90 95 cag ctc gac cac ccg gtg gtg cgc tcg gcc gcc gtg cgc ccc gtc tgc 336Gln Leu Asp His Pro Val Val Arg Ser Ala Ala Val Arg Pro Val Cys 100 105 110 ctg ccc gcg cgc tcc cac ttc ttc gag ccc ggc ctg cac tgc tgg att 384 Leu Pro Ala Arg Ser His Phe Phe Glu Pro Gly Leu His Cys Trp Ile 115 120 125 acg ggc tgg ggc gcc ttg cgc gag ggc ggc ccc atc agc aac gct ctg 432Thr Gly Trp Gly Ala Leu Arg Glu Gly Gly Pro Ile Ser Asn Ala Leu 130 135 140 cag aaa gtg gat gtg cag ttg atc cca cag gac ctg tgc agc gag gtc 480Gln Lys Val Asp Val Gln Leu Ile Pro Gln Asp Leu Cys Ser Glu Val 145 150 155 160 tat cgc tac cag gtg acg cca cgc atg ctg tgt gcc ggc tac cgc aag 528 Tyr Arg Tyr Gln Val Thr Pro Arg Met Leu Cys Ala Gly Tyr Arg Lys 165 170 175 ggc aag aag gat gcc tgt cag ggt gac tca ggt ggt ccg ctg gtg tgc 576 Gly Lys Lys Asp Ala Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Cys 180 185 190 aag gca ctc agt ggc cgc tgg ttc ctg gcg ggg ctg gtc agc tgg ggc 624 Lys Ala Leu Ser Gly Arg Trp Phe Leu Ala Gly Leu Val Ser Trp Gly 195 200 205 ctg ggc tgt ggc cgg cct aac tac ttc ggc gtc tac acc cgc atc aca 672 Leu Gly Cys Gly Arg Pro Asn Tyr Phe Gly Val Tyr Thr Arg Ile Thr 210 215 220 ggt gtg atc agc tgg atc cag caa gtg gtg acc tga 708 Gly Val Ile Ser Trp Ile Gln Gln Val Val Thr * 225 230 235 <210> SEQ ID NO 6 <211> LENGTH: 235 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 6 Ile Val Gly Gly Ala Val Ser Ser Glu Gly Glu Trp Pro Trp Gln Ala 1 5 10 15 Ser Leu Gln Val Arg Gly Arg His Ile Cys Gly Gly Ala Leu Ile Ala 20 25 30 Asp Arg Trp Val Ile Thr Ala Ala His Cys Phe Gln Glu Asp Ser Met 35 40 45 Ala Ser Thr Val Leu Trp Thr Val Phe Leu Gly Lys Val Trp Gln Asn 50 55 60 Ser Arg Trp Pro Gly Glu Val Ser Phe Lys Val Ser Arg Leu Leu Leu 65 70 75 80 His Pro Tyr His Glu Glu Asp Ser His Asp Tyr Asp Val Ala Leu Leu 85 90 95 Gln Leu Asp His Pro Val Val Arg Ser Ala Ala Val Arg Pro Val Cys 100 105 110 Leu Pro Ala Arg Ser His Phe Phe Glu Pro Gly Leu His Cys Trp Ile 115 120 125 Thr Gly Trp Gly Ala Leu Arg Glu Gly Gly Pro Ile Ser Asn Ala Leu 130 135 140 Gln Lys Val Asp Val Gln Leu Ile Pro Gln Asp Leu Cys Ser Glu Val 145 150 155 160 Tyr Arg Tyr Gln Val Thr Pro Arg Met Leu Cys Ala Gly Tyr Arg Lys 165 170 175 Gly Lys Lys Asp Ala Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Cys 180 185 190 Lys Ala Leu Ser Gly Arg Trp Phe Leu Ala Gly Leu Val Ser Trp Gly 195 200 205 Leu Gly Cys Gly Arg Pro Asn Tyr Phe Gly Val Tyr Thr Arg Ile Thr 210 215 220 Gly Val Ile Ser Trp Ile Gln Gln Val Val Thr 225 230 235 <210> SEQ ID NO 7 <211> LENGTH: 3104 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (33)...(2441) <223> OTHER INFORMATION: Nucleic acid encoding MTSP4-L (long form) splice variant <400> SEQUENCE: 7 tcatcggcca gagggtgatc agtgagcaga ag atg ccc gtg gcc gag gcc ccc 53 Met Pro Val Ala Glu Ala Pro 1 5 cag gtg gct ggc ggg cag ggg gac gga ggt gat ggc gag gaa gcg gag 101 Gln Val Ala Gly Gly Gln Gly Asp Gly Gly Asp Gly Glu Glu Ala Glu 10 15 20 ccg gag ggg atg ttc aag gcc tgt gag gac tcc aag aga aaa gcc cgg 149 Pro Glu Gly Met Phe Lys Ala Cys Glu Asp Ser Lys Arg Lys Ala Arg 25 30 35 ggc tac ctc cgc ctg gtg ccc ctg ttt gtg ctg ctg gcc ctg ctc gtg 197 Gly Tyr Leu Arg Leu Val Pro Leu Phe Val Leu Leu Ala Leu Leu Val 40 45 50 55 ctg gct tcg gcg ggg gtg cta ctc tgg tat ttc cta ggg tac aag gcg 245 Leu Ala Ser Ala Gly Val Leu Leu Trp Tyr Phe Leu Gly Tyr Lys Ala 60 65 70 gag gtg atg gtc agc cag gtg tac tca ggc agt ctg cgt gta ctc aat 293Glu Val Met Val Ser Gln Val Tyr Ser Gly Ser Leu Arg Val Leu Asn 75 80 85 cgc cac ttc tcc cag gat ctt acc cgc cgg gaa tct agt gcc ttc cgc 341Arg His Phe Ser Gln Asp Leu Thr Arg Arg Glu Ser Ser Ala Phe Arg 90 95 100 agt gaa acc gcc aaa gcc cag aag atg ctc aag gag ctc atc acc agc 389Ser Glu Thr Ala Lys Ala Gln Lys Met Leu Lys Glu Leu Ile Thr Ser 105 110 115 acc cgc ctg gga act tac tac aac tcc agc tcc gtc tat tcc ttt ggg 437Thr Arg Leu Gly Thr Tyr Tyr Asn Ser Ser Ser Val Tyr Ser Phe Gly 120 125 130 135 gag gga ccc ctc acc tgc ttc ttc tgg ttc att ctc caa atc ccc gag 485 Glu Gly Pro Leu Thr Cys Phe Phe Trp Phe Ile Leu Gln Ile Pro Glu 140 145 150 cac cgc cgg ctg atg ctg agc ccc gag gtg gtg cag gca ctg ctg gtg 533His Arg Arg Leu Met Leu Ser Pro Glu Val Val Gln Ala Leu Leu Val 155 160 165 gag gag ctg ctg tcc aca gtc aac agc tcg gct gcc gtc ccc tac agg 581 Glu Glu Leu Leu Ser Thr Val Asn Ser Ser Ala Ala Val Pro Tyr Arg 170 175 180 gcc gag tac gaa gtg gac ccc gag ggc cta gtg atc ctg gaa gcc agt 629 Ala Glu Tyr Glu Val Asp Pro Glu Gly Leu Val Ile Leu Glu Ala Ser 185 190 195 gtg aaa gac ata gct gca ttg aat tcc acg ctg ggt tgt tac cgc tac 677 Val Lys Asp Ile Ala Ala Leu Asn Ser Thr Leu Gly Cys Tyr Arg Tyr 200 205 210 215 agc tac gtg ggc cag ggc cag gtc ctc cgg ctg aag ggg cct gac cac 725 Ser Tyr Val Gly Gln Gly Gln Val Leu Arg Leu Lys Gly Pro Asp His 220 225 230 ctg gcc tcc agc tgc ctg tgg cac ctg cag ggc ccc aag gac ctc atg 773 Leu Ala Ser Ser Cys Leu Trp His Leu Gln Gly Pro Lys Asp Leu Met 235 240 245 ctc aaa ctc cgg ctg gag tgg acg ctg gca gag tgc cgg gac cga ctg 821 Leu Lys Leu Arg Leu Glu Trp Thr Leu Ala Glu Cys Arg Asp Arg Leu 250 255 260 gcc atg tat gac gtg gcc ggg ccc ctg gag aag agg ctc atc acc tcg 869 Ala Met Tyr Asp Val Ala Gly Pro Leu Glu Lys Arg Leu Ile Thr Ser 265 270 275 gtg tac ggc tgc agc cgc cag gag ccc gtg gtg gag gtt ctg gcg tcg 917 Val Tyr Gly Cys Ser Arg Gln Glu Pro Val Val Glu Val Leu Ala Ser 280 285 290 295 ggg gcc atc atg gcg gtc gtc tgg aag aag ggc ctg cac agc tac tac 965 Gly Ala Ile Met Ala Val Val Trp Lys Lys Gly Leu His Ser Tyr Tyr 300 305 310 gac ccc ttc gtg ctc tcc gtg cag ccg gtg gtc ttc cag gcc tgt gaa 1013 Asp Pro Phe Val Leu Ser Val Gln Pro Val Val Phe Gln Ala Cys Glu 315 320 325 gtg aac ctg acg ctg gac aac agg ctc gac tcc cag ggc gtc ctc agc 1061 Val Asn Leu Thr Leu Asp Asn Arg Leu Asp Ser Gln Gly Val Leu Ser 330 335 340 acc ccg tac ttc ccc agc tac tac tcg ccc caa acc cac tgc tcc tgg 1109 Thr Pro Tyr Phe Pro Ser Tyr Tyr Ser Pro Gln Thr His Cys Ser Trp 345 350 355 cac ctc acg gtg ccc tct ctg gac tac ggc ttg gcc ctc tgg ttt gat 1157 His Leu Thr Val Pro Ser Leu Asp Tyr Gly Leu Ala Leu Trp Phe Asp 360 365 370 375 gcc tat gca ctg agg agg cag aag tat gat ttg ccg tgc acc cag ggc 1205 Ala Tyr Ala Leu Arg Arg Gln Lys Tyr Asp Leu Pro Cys Thr Gln Gly 380 385 390 cag tgg acg atc cag aac agg agg ctg tgt ggc ttg cgc atc ctg cag 1253 Gln Trp Thr Ile Gln Asn Arg Arg Leu Cys Gly Leu Arg Ile Leu Gln 395 400 405 ccc tac gcc gag agg atc ccc gtg gtg gcc acg gcc ggg atc acc atc 1301 Pro Tyr Ala Glu Arg Ile Pro Val Val Ala Thr Ala Gly Ile Thr Ile 410 415 420 aac ttc acc tcc cag atc tcc ctc acc ggg ccc ggt gtg cgg gtg cac 1349 Asn Phe Thr Ser Gln Ile Ser Leu Thr Gly Pro Gly Val Arg Val His 425 430 435 tat ggc ttg tac aac cag tcg gac ccc tgc cct gga gag ttc ctc tgt 1397 Tyr Gly Leu Tyr Asn Gln Ser Asp Pro Cys Pro Gly Glu Phe Leu Cys 440 445 450 455 tct gtg aat gga ctc tgt gtc cct gcc tgt gat ggg gtc aag gac tgc 1445 Ser Val Asn Gly Leu Cys Val Pro Ala Cys Asp Gly Val Lys Asp Cys 460 465 470 ccc aac ggc ctg gat gag aga aac tgc gtt tgc aga gcc aca ttc cag 1493 Pro Asn Gly Leu Asp Glu Arg Asn Cys Val Cys Arg Ala Thr Phe Gln 475 480 485 tgc aaa gag gac agc aca tgc atc tca ctg ccc aag gtc tgt gat ggg 1541 Cys Lys Glu Asp Ser Thr Cys Ile Ser Leu Pro Lys Val Cys Asp Gly 490 495 500 cag cct gat tgt ctc aac ggc agc gac gaa gag cag tgc cag gaa ggg 1589 Gln Pro Asp Cys Leu Asn Gly Ser Asp Glu Glu Gln Cys Gln Glu Gly 505 510 515 gtg cca tgt ggg aca ttc acc ttc cag tgt gag gac cgg agc tgc gtg 1637 Val Pro Cys Gly Thr Phe Thr Phe Gln Cys Glu Asp Arg Ser Cys Val 520 525 530 535 aag aag ccc aac ccg cag tgt gat ggg cgg ccc gac tgc agg gac ggc 1685 Lys Lys Pro Asn Pro Gln Cys Asp Gly Arg Pro Asp Cys Arg Asp Gly 540 545 550 tcg gat gag gag cac tgt gaa tgt ggc ctc cag ggc ccc tcc agc cgc 1733 Ser Asp Glu Glu His Cys Glu Cys Gly Leu Gln Gly Pro Ser Ser Arg 555 560 565 att gtt ggt gga gct gtg tcc tcc gag ggt gag tgg cca tgg cag gcc 1781 Ile Val Gly Gly Ala Val Ser Ser Glu Gly Glu Trp Pro Trp Gln Ala 570 575 580 agc ctc cag gtt cgg ggt cga cac atc tgt ggg ggg gcc ctc atc gct 1829 Ser Leu Gln Val Arg Gly Arg His Ile Cys Gly Gly Ala Leu Ile Ala 585 590 595 gac cgc tgg gtg ata aca gct gcc cac tgc ttc cag gag gac agc atg 1877 Asp Arg Trp Val Ile Thr Ala Ala His Cys Phe Gln Glu Asp Ser Met 600 605 610 615 gcc tcc acg gtg ctg tgg acc gtg ttc ctg ggc aag gtg tgg cag aac 1925 Ala Ser Thr Val Leu Trp Thr Val Phe Leu Gly Lys Val Trp Gln Asn 620 625 630 tcg cgc tgg cct gga gag gtg tcc ttc aag gtg agc cgc ctg ctc ctg 1973 Ser Arg Trp Pro Gly Glu Val Ser Phe Lys Val Ser Arg Leu Leu Leu 635 640 645 cac ccg tac cac gaa gag gac agc cat gac tac gac gtg gcg ctg ctg 2021 His Pro Tyr His Glu Glu Asp Ser His Asp Tyr Asp Val Ala Leu Leu 650 655 660 cag ctc gac cac ccg gtg gtg cgc tcg gcc gcc gtg cgc ccc gtc tgc 2069 Gln Leu Asp His Pro Val Val Arg Ser Ala Ala Val Arg Pro Val Cys 665 670 675 ctg ccc gcg cgc tcc cac ttc ttc gag ccc ggc ctg cac tgc tgg att 2117 Leu Pro Ala Arg Ser His Phe Phe Glu Pro Gly Leu His Cys Trp Ile 680 685 690 695 acg ggc tgg ggc gcc ttg cgc gag ggc ggc ccc atc agc aac gct ctg 2165 Thr Gly Trp Gly Ala Leu Arg Glu Gly Gly Pro Ile Ser Asn Ala Leu 700 705 710 cag aaa gtg gat gtg cag ttg atc cca cag gac ctg tgc agc gag gtc 2213 Gln Lys Val Asp Val Gln Leu Ile Pro Gln Asp Leu Cys Ser Glu Val 715 720 725 tat cgc tac cag gtg acg cca cgc atg ctg tgt gcc ggc tac cgc aag 2261 Tyr Arg Tyr Gln Val Thr Pro Arg Met Leu Cys Ala Gly Tyr Arg Lys 730 735 740 ggc aag aag gat gcc tgt cag ggt gac tca ggt ggt ccg ctg gtg tgc 2309 Gly Lys Lys Asp Ala Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Cys 745 750 755 aag gca ctc agt ggc cgc tgg ttc ctg gcg ggg ctg gtc agc tgg ggc 2357 Lys Ala Leu Ser Gly Arg Trp Phe Leu Ala Gly Leu Val Ser Trp Gly 760 765 770 775 ctg ggc tgt ggc cgg cct aac tac ttc ggc gtc tac acc cgc atc aca 2405 Leu Gly Cys Gly Arg Pro Asn Tyr Phe Gly Val Tyr Thr Arg Ile Thr 780 785 790 ggt gtg atc agc tgg atc cag caa gtg gtg acc tga ggaactgccc 2451 Gly Val Ile Ser Trp Ile Gln Gln Val Val Thr * 795 800 ccctgcaaag cagggcccac ctcctggact cagagagccc agggcaactg ccaagcaggg 2511 ggacaagtat tctggcgggg ggtgggggag agagcaggcc ctgtggtggc aggaggggca 2571 tcttgtttcg tccctgatgt ctgtccagta tggcaggagg atgagaagtg ccagcagttg 2631 ggggtcaaga cgtcccttga ggacccaggc ccacacccag cccttttgcc tcccaattct 2691 ctctcctccg tccccttcct ccactgctgc ctaatgcaag gcagtggctc agcagcaaga 2751 atgctggttc tacatcccga ggagtgtctg aggtgcgccc cactctgtac agaggctgtt 2811 tgggcagcct tgcctccaga gagcagattc cagcttcgga agcccctggt ctaacttggg 2871 atctgggaat ggaaggtgct cccatcggag gggaccctca gagccctgga gactgccagg 2931 tgggcctgct gccactgtaa gccaaaaggt ggggaagtcc tgactccagg gtccttgccc 2991 cacccctgcc tgccacctgg gccctcacag cccagaccct cactgggagg tgagctcagc 3051 tgccctttgg aataaagctg cctgatgcaa aaaaaaaaaa aaaaaaaaaa aaa 3104 <210> SEQ ID NO 8 <211> LENGTH: 802 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 8 Met Pro Val Ala Glu Ala Pro Gln Val Ala Gly Gly Gln Gly Asp Gly 1 5 10 15 Gly Asp Gly Glu Glu Ala Glu Pro Glu Gly Met Phe Lys Ala Cys Glu 20 25 30 Asp Ser Lys Arg Lys Ala Arg Gly Tyr Leu Arg Leu Val Pro Leu Phe 35 40 45 Val Leu Leu Ala Leu Leu Val Leu Ala Ser Ala Gly Val Leu Leu Trp 50 55 60 Tyr Phe Leu Gly Tyr Lys Ala Glu Val Met Val Ser Gln Val Tyr Ser 65 70 75 80 Gly Ser Leu Arg Val Leu Asn Arg His Phe Ser Gln Asp Leu Thr Arg 85 90 95 Arg Glu Ser Ser Ala Phe Arg Ser Glu Thr Ala Lys Ala Gln Lys Met 100 105 110 Leu Lys Glu Leu Ile Thr Ser Thr Arg Leu Gly Thr Tyr Tyr Asn Ser 115 120 125 Ser Ser Val Tyr Ser Phe Gly Glu Gly Pro Leu Thr Cys Phe Phe Trp 130 135 140 Phe Ile Leu Gln Ile Pro Glu His Arg Arg Leu Met Leu Ser Pro Glu 145 150 155 160 Val Val Gln Ala Leu Leu Val Glu Glu Leu Leu Ser Thr Val Asn Ser 165 170 175 Ser Ala Ala Val Pro Tyr Arg Ala Glu Tyr Glu Val Asp Pro Glu Gly 180 185 190 Leu Val Ile Leu Glu Ala Ser Val Lys Asp Ile Ala Ala Leu Asn Ser 195 200 205 Thr Leu Gly Cys Tyr Arg Tyr Ser Tyr Val Gly Gln Gly Gln Val Leu 210 215 220 Arg Leu Lys Gly Pro Asp His Leu Ala Ser Ser Cys Leu Trp His Leu 225 230 235 240 Gln Gly Pro Lys Asp Leu Met Leu Lys Leu Arg Leu Glu Trp Thr Leu 245 250 255 Ala Glu Cys Arg Asp Arg Leu Ala Met Tyr Asp Val Ala Gly Pro Leu 260 265 270 Glu Lys Arg Leu Ile Thr Ser Val Tyr Gly Cys Ser Arg Gln Glu Pro 275 280 285 Val Val Glu Val Leu Ala Ser Gly Ala Ile Met Ala Val Val Trp Lys 290 295 300 Lys Gly Leu His Ser Tyr Tyr Asp Pro Phe Val Leu Ser Val Gln Pro 305 310 315 320 Val Val Phe Gln Ala Cys Glu Val Asn Leu Thr Leu Asp Asn Arg Leu 325 330 335 Asp Ser Gln Gly Val Leu Ser Thr Pro Tyr Phe Pro Ser Tyr Tyr Ser 340 345 350 Pro Gln Thr His Cys Ser Trp His Leu Thr Val Pro Ser Leu Asp Tyr 355 360 365 Gly Leu Ala Leu Trp Phe Asp Ala Tyr Ala Leu Arg Arg Gln Lys Tyr 370 375 380 Asp Leu Pro Cys Thr Gln Gly Gln Trp Thr Ile Gln Asn Arg Arg Leu 385 390 395 400 Cys Gly Leu Arg Ile Leu Gln Pro Tyr Ala Glu Arg Ile Pro Val Val 405 410 415 Ala Thr Ala Gly Ile Thr Ile Asn Phe Thr Ser Gln Ile Ser Leu Thr 420 425 430 Gly Pro Gly Val Arg Val His Tyr Gly Leu Tyr Asn Gln Ser Asp Pro 435 440 445 Cys Pro Gly Glu Phe Leu Cys Ser Val Asn Gly Leu Cys Val Pro Ala 450 455 460 Cys Asp Gly Val Lys Asp Cys Pro Asn Gly Leu Asp Glu Arg Asn Cys 465 470 475 480 Val Cys Arg Ala Thr Phe Gln Cys Lys Glu Asp Ser Thr Cys Ile Ser 485 490 495 Leu Pro Lys Val Cys Asp Gly Gln Pro Asp Cys Leu Asn Gly Ser Asp 500 505 510 Glu Glu Gln Cys Gln Glu Gly Val Pro Cys Gly Thr Phe Thr Phe Gln 515 520 525 Cys Glu Asp Arg Ser Cys Val Lys Lys Pro Asn Pro Gln Cys Asp Gly 530 535 540 Arg Pro Asp Cys Arg Asp Gly Ser Asp Glu Glu His Cys Glu Cys Gly 545 550 555 560 Leu Gln Gly Pro Ser Ser Arg Ile Val Gly Gly Ala Val Ser Ser Glu 565 570 575 Gly Glu Trp Pro Trp Gln Ala Ser Leu Gln Val Arg Gly Arg His Ile 580 585 590 Cys Gly Gly Ala Leu Ile Ala Asp Arg Trp Val Ile Thr Ala Ala His 595 600 605 Cys Phe Gln Glu Asp Ser Met Ala Ser Thr Val Leu Trp Thr Val Phe 610 615 620 Leu Gly Lys Val Trp Gln Asn Ser Arg Trp Pro Gly Glu Val Ser Phe 625 630 635 640 Lys Val Ser Arg Leu Leu Leu His Pro Tyr His Glu Glu Asp Ser His 645 650 655 Asp Tyr Asp Val Ala Leu Leu Gln Leu Asp His Pro Val Val Arg Ser 660 665 670 Ala Ala Val Arg Pro Val Cys Leu Pro Ala Arg Ser His Phe Phe Glu 675 680 685 Pro Gly Leu His Cys Trp Ile Thr Gly Trp Gly Ala Leu Arg Glu Gly 690 695 700 Gly Pro Ile Ser Asn Ala Leu Gln Lys Val Asp Val Gln Leu Ile Pro 705 710 715 720 Gln Asp Leu Cys Ser Glu Val Tyr Arg Tyr Gln Val Thr Pro Arg Met 725 730 735 Leu Cys Ala Gly Tyr Arg Lys Gly Lys Lys Asp Ala Cys Gln Gly Asp 740 745 750 Ser Gly Gly Pro Leu Val Cys Lys Ala Leu Ser Gly Arg Trp Phe Leu 755 760 765 Ala Gly Leu Val Ser Trp Gly Leu Gly Cys Gly Arg Pro Asn Tyr Phe 770 775 780 Gly Val Tyr Thr Arg Ile Thr Gly Val Ile Ser Trp Ile Gln Gln Val 785 790 795 800 Val Thr <210> SEQ ID NO 9 <211> LENGTH: 2672 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (33)...(2009) <223> OTHER INFORMATION: cDNA encoding: MTSP4-S (short form) splice variant <400> SEQUENCE: 9 tcatcggcca gagggtgatc agtgagcaga ag atg ccc gtg gcc gag gcc ccc 53 Met Pro Val Ala Glu Ala Pro 1 5 cag gtg gct ggc ggg cag ggg gac gga ggt gat ggc gag gaa gcg gag 101 Gln Val Ala Gly Gly Gln Gly Asp Gly Gly Asp Gly Glu Glu Ala Glu 10 15 20 ccg gag ggg atg ttc aag gcc tgt gag gac tcc aag aga aaa gcc cgg 149 Pro Glu Gly Met Phe Lys Ala Cys Glu Asp Ser Lys Arg Lys Ala Arg 25 30 35 ggc tac ctc cgc ctg gtg ccc ctg ttt gtg ctg ctg gcc ctg ctc gtg 197 Gly Tyr Leu Arg Leu Val Pro Leu Phe Val Leu Leu Ala Leu Leu Val 40 45 50 55 ctg gct tcg gcg ggg gtg cta ctc tgg tat ttc cta ggg tac aag gcg 245 Leu Ala Ser Ala Gly Val Leu Leu Trp Tyr Phe Leu Gly Tyr Lys Ala 60 65 70 gag gtg atg gtc agc cag gtg tac tca ggc agt ctg cgt gta ctc aat 293Glu Val Met Val Ser Gln Val Tyr Ser Gly Ser Leu Arg Val Leu Asn 75 80 85 cgc cac ttc tcc cag gat ctt acc cgc cgg gaa tct agt gcc ttc cgc 341Arg His Phe Ser Gln Asp Leu Thr Arg Arg Glu Ser Ser Ala Phe Arg 90 95 100 agt gaa acc gcc aaa gcc cag aag atg ctc aag gag ctc atc acc agc 389Ser Glu Thr Ala Lys Ala Gln Lys Met Leu Lys Glu Leu Ile Thr Ser 105 110 115 acc cgc ctg gga act tac tac aac tcc agc tcc gtc tat tcc ttt ggg 437Thr Arg Leu Gly Thr Tyr Tyr Asn Ser Ser Ser Val Tyr Ser Phe Gly 120 125 130 135 gtg tac ggc tgc agc cgc cag gag ccc gtg gtg gag gtt ctg gcg tcg 485Val Tyr Gly Cys Ser Arg Gln Glu Pro Val Val Glu Val Leu Ala Ser 140 145 150 ggg gcc atc atg gcg gtc gtc tgg aag aag ggc ctg cac agc tac tac 533Gly Ala Ile Met Ala Val Val Trp Lys Lys Gly Leu His Ser Tyr Tyr 155 160 165 gac ccc ttc gtg ctc tcc gtg cag ccg gtg gtc ttc cag gcc tgt gaa 581 Asp Pro Phe Val Leu Ser Val Gln Pro Val Val Phe Gln Ala Cys Glu 170 175 180 gtg aac ctg acg ctg gac aac agg ctc gac tcc cag ggc gtc ctc agc 629 Val Asn Leu Thr Leu Asp Asn Arg Leu Asp Ser Gln Gly Val Leu Ser 185 190 195 acc ccg tac ttc ccc agc tac tac tcg ccc caa acc cac tgc tcc tgg 677 Thr Pro Tyr Phe Pro Ser Tyr Tyr Ser Pro Gln Thr His Cys Ser Trp 200 205 210 215 cac ctc acg gtg ccc tct ctg gac tac ggc ttg gcc ctc tgg ttt gat 725 His Leu Thr Val Pro Ser Leu Asp Tyr Gly Leu Ala Leu Trp Phe Asp 220 225 230 gcc tat gca ctg agg agg cag aag tat gat ttg ccg tgc acc cag ggc 773 Ala Tyr Ala Leu Arg Arg Gln Lys Tyr Asp Leu Pro Cys Thr Gln Gly 235 240 245 cag tgg acg atc cag aac agg agg ctg tgt ggc ttg cgc atc ctg cag 821 Gln Trp Thr Ile Gln Asn Arg Arg Leu Cys Gly Leu Arg Ile Leu Gln 250 255 260 ccc tac gcc gag agg atc ccc gtg gtg gcc acg gcc ggg atc acc atc 869 Pro Tyr Ala Glu Arg Ile Pro Val Val Ala Thr Ala Gly Ile Thr Ile 265 270 275 aac ttc acc tcc cag atc tcc ctc acc ggg ccc ggt gtg cgg gtg cac 917 Asn Phe Thr Ser Gln Ile Ser Leu Thr Gly Pro Gly Val Arg Val His 280 285 290 295 tat ggc ttg tac aac cag tcg gac ccc tgc cct gga gag ttc ctc tgt 965 Tyr Gly Leu Tyr Asn Gln Ser Asp Pro Cys Pro Gly Glu Phe Leu Cys 300 305 310 tct gtg aat gga ctc tgt gtc cct gcc tgt gat ggg gtc aag gac tgc 1013 Ser Val Asn Gly Leu Cys Val Pro Ala Cys Asp Gly Val Lys Asp Cys 315 320 325 ccc aac ggc ctg gat gag aga aac tgc gtt tgc aga gcc aca ttc cag 1061 Pro Asn Gly Leu Asp Glu Arg Asn Cys Val Cys Arg Ala Thr Phe Gln 330 335 340 tgc aaa gag gac agc aca tgc atc tca ctg ccc aag gtc tgt gat ggg 1109 Cys Lys Glu Asp Ser Thr Cys Ile Ser Leu Pro Lys Val Cys Asp Gly 345 350 355 cag cct gat tgt ctc aac ggc agc gac gaa gag cag tgc cag gaa ggg 1157 Gln Pro Asp Cys Leu Asn Gly Ser Asp Glu Glu Gln Cys Gln Glu Gly 360 365 370 375 gtg cca tgt ggg aca ttc acc ttc cag tgt gag gac cgg agc tgc gtg 1205 Val Pro Cys Gly Thr Phe Thr Phe Gln Cys Glu Asp Arg Ser Cys Val 380 385 390 aag aag ccc aac ccg cag tgt gat ggg cgg ccc gac tgc agg gac ggc 1253 Lys Lys Pro Asn Pro Gln Cys Asp Gly Arg Pro Asp Cys Arg Asp Gly 395 400 405 tcg gat gag gag cac tgt gaa tgt ggc ctc cag ggc ccc tcc agc cgc 1301 Ser Asp Glu Glu His Cys Glu Cys Gly Leu Gln Gly Pro Ser Ser Arg 410 415 420 att gtt ggt gga gct gtg tcc tcc gag ggt gag tgg cca tgg cag gcc 1349 Ile Val Gly Gly Ala Val Ser Ser Glu Gly Glu Trp Pro Trp Gln Ala 425 430 435 agc ctc cag gtt cgg ggt cga cac atc tgt ggg ggg gcc ctc atc gct 1397 Ser Leu Gln Val Arg Gly Arg His Ile Cys Gly Gly Ala Leu Ile Ala 440 445 450 455 gac cgc tgg gtg ata aca gct gcc cac tgc ttc cag gag gac agc atg 1445 Asp Arg Trp Val Ile Thr Ala Ala His Cys Phe Gln Glu Asp Ser Met 460 465 470 gcc tcc acg gtg ctg tgg acc gtg ttc ctg ggc aag gtg tgg cag aac 1493 Ala Ser Thr Val Leu Trp Thr Val Phe Leu Gly Lys Val Trp Gln Asn 475 480 485 tcg cgc tgg cct gga gag gtg tcc ttc aag gtg agc cgc ctg ctc ctg 1541 Ser Arg Trp Pro Gly Glu Val Ser Phe Lys Val Ser Arg Leu Leu Leu 490 495 500 cac ccg tac cac gaa gag gac agc cat gac tac gac gtg gcg ctg ctg 1589 His Pro Tyr His Glu Glu Asp Ser His Asp Tyr Asp Val Ala Leu Leu 505 510 515 cag ctc gac cac ccg gtg gtg cgc tcg gcc gcc gtg cgc ccc gtc tgc 1637 Gln Leu Asp His Pro Val Val Arg Ser Ala Ala Val Arg Pro Val Cys 520 525 530 535 ctg ccc gcg cgc tcc cac ttc ttc gag ccc ggc ctg cac tgc tgg att 1685 Leu Pro Ala Arg Ser His Phe Phe Glu Pro Gly Leu His Cys Trp Ile 540 545 550 acg ggc tgg ggc gcc ttg cgc gag ggc ggc ccc atc agc aac gct ctg 1733 Thr Gly Trp Gly Ala Leu Arg Glu Gly Gly Pro Ile Ser Asn Ala Leu 555 560 565 cag aaa gtg gat gtg cag ttg atc cca cag gac ctg tgc agc gag gtc 1781 Gln Lys Val Asp Val Gln Leu Ile Pro Gln Asp Leu Cys Ser Glu Val 570 575 580 tat cgc tac cag gtg acg cca cgc atg ctg tgt gcc ggc tac cgc aag 1829 Tyr Arg Tyr Gln Val Thr Pro Arg Met Leu Cys Ala Gly Tyr Arg Lys 585 590 595 ggc aag aag gat gcc tgt cag ggt gac tca ggt ggt ccg ctg gtg tgc 1877 Gly Lys Lys Asp Ala Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Cys 600 605 610 615 aag gca ctc agt ggc cgc tgg ttc ctg gcg ggg ctg gtc agc tgg ggc 1925 Lys Ala Leu Ser Gly Arg Trp Phe Leu Ala Gly Leu Val Ser Trp Gly 620 625 630 ctg ggc tgt ggc cgg cct aac tac ttc ggc gtc tac acc cgc atc aca 1973 Leu Gly Cys Gly Arg Pro Asn Tyr Phe Gly Val Tyr Thr Arg Ile Thr 635 640 645 ggt gtg atc agc tgg atc cag caa gtg gtg acc tga ggaactgccc 2019 Gly Val Ile Ser Trp Ile Gln Gln Val Val Thr * 650 655 ccctgcaaag cagggcccac ctcctggact cagagagccc agggcaactg ccaagcaggg 2079 ggacaagtat tctggcgggg ggtgggggag agagcaggcc ctgtggtggc aggaggggca 2139 tcttgtttcg tccctgatgt ctgtccagta tggcaggagg atgagaagtg ccagcagttg 2199 ggggtcaaga cgtcccttga ggacccaggc ccacacccag cccttttgcc tcccaattct 2259 ctctcctccg tccccttcct ccactgctgc ctaatgcaag gcagtggctc agcagcaaga 2319 atgctggttc tacatcccga ggagtgtctg aggtgcgccc cactctgtac agaggctgtt 2379 tgggcagcct tgcctccaga gagcagattc cagcttcgga agcccctggt ctaacttggg 2439 atctgggaat ggaaggtgct cccatcggag gggaccctca gagccctgga gactgccagg 2499 tgggcctgct gccactgtaa gccaaaaggt ggggaagtcc tgactccagg gtccttgccc 2559 cacccctgcc tgccacctgg gccctcacag cccagaccct cactgggagg tgagctcagc 2619 tgccctttgg aataaagctg cctgatgcaa aaaaaaaaaa aaaaaaaaaa aaa 2672 <210> SEQ ID NO 10 <211> LENGTH: 658 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 10 Met Pro Val Ala Glu Ala Pro Gln Val Ala Gly Gly Gln Gly Asp Gly 1 5 10 15 Gly Asp Gly Glu Glu Ala Glu Pro Glu Gly Met Phe Lys Ala Cys Glu 20 25 30 Asp Ser Lys Arg Lys Ala Arg Gly Tyr Leu Arg Leu Val Pro Leu Phe 35 40 45 Val Leu Leu Ala Leu Leu Val Leu Ala Ser Ala Gly Val Leu Leu Trp 50 55 60 Tyr Phe Leu Gly Tyr Lys Ala Glu Val Met Val Ser Gln Val Tyr Ser 65 70 75 80 Gly Ser Leu Arg Val Leu Asn Arg His Phe Ser Gln Asp Leu Thr Arg 85 90 95 Arg Glu Ser Ser Ala Phe Arg Ser Glu Thr Ala Lys Ala Gln Lys Met 100 105 110 Leu Lys Glu Leu Ile Thr Ser Thr Arg Leu Gly Thr Tyr Tyr Asn Ser 115 120 125 Ser Ser Val Tyr Ser Phe Gly Val Tyr Gly Cys Ser Arg Gln Glu Pro 130 135 140 Val Val Glu Val Leu Ala Ser Gly Ala Ile Met Ala Val Val Trp Lys 145 150 155 160 Lys Gly Leu His Ser Tyr Tyr Asp Pro Phe Val Leu Ser Val Gln Pro 165 170 175 Val Val Phe Gln Ala Cys Glu Val Asn Leu Thr Leu Asp Asn Arg Leu 180 185 190 Asp Ser Gln Gly Val Leu Ser Thr Pro Tyr Phe Pro Ser Tyr Tyr Ser 195 200 205 Pro Gln Thr His Cys Ser Trp His Leu Thr Val Pro Ser Leu Asp Tyr 210 215 220 Gly Leu Ala Leu Trp Phe Asp Ala Tyr Ala Leu Arg Arg Gln Lys Tyr 225 230 235 240 Asp Leu Pro Cys Thr Gln Gly Gln Trp Thr Ile Gln Asn Arg Arg Leu 245 250 255 Cys Gly Leu Arg Ile Leu Gln Pro Tyr Ala Glu Arg Ile Pro Val Val 260 265 270 Ala Thr Ala Gly Ile Thr Ile Asn Phe Thr Ser Gln Ile Ser Leu Thr 275 280 285 Gly Pro Gly Val Arg Val His Tyr Gly Leu Tyr Asn Gln Ser Asp Pro 290 295 300 Cys Pro Gly Glu Phe Leu Cys Ser Val Asn Gly Leu Cys Val Pro Ala 305 310 315 320 Cys Asp Gly Val Lys Asp Cys Pro Asn Gly Leu Asp Glu Arg Asn Cys 325 330 335 Val Cys Arg Ala Thr Phe Gln Cys Lys Glu Asp Ser Thr Cys Ile Ser 340 345 350 Leu Pro Lys Val Cys Asp Gly Gln Pro Asp Cys Leu Asn Gly Ser Asp 355 360 365 Glu Glu Gln Cys Gln Glu Gly Val Pro Cys Gly Thr Phe Thr Phe Gln 370 375 380 Cys Glu Asp Arg Ser Cys Val Lys Lys Pro Asn Pro Gln Cys Asp Gly 385 390 395 400 Arg Pro Asp Cys Arg Asp Gly Ser Asp Glu Glu His Cys Glu Cys Gly 405 410 415 Leu Gln Gly Pro Ser Ser Arg Ile Val Gly Gly Ala Val Ser Ser Glu 420 425 430 Gly Glu Trp Pro Trp Gln Ala Ser Leu Gln Val Arg Gly Arg His Ile 435 440 445 Cys Gly Gly Ala Leu Ile Ala Asp Arg Trp Val Ile Thr Ala Ala His 450 455 460 Cys Phe Gln Glu Asp Ser Met Ala Ser Thr Val Leu Trp Thr Val Phe 465 470 475 480 Leu Gly Lys Val Trp Gln Asn Ser Arg Trp Pro Gly Glu Val Ser Phe 485 490 495 Lys Val Ser Arg Leu Leu Leu His Pro Tyr His Glu Glu Asp Ser His 500 505 510 Asp Tyr Asp Val Ala Leu Leu Gln Leu Asp His Pro Val Val Arg Ser 515 520 525 Ala Ala Val Arg Pro Val Cys Leu Pro Ala Arg Ser His Phe Phe Glu 530 535 540 Pro Gly Leu His Cys Trp Ile Thr Gly Trp Gly Ala Leu Arg Glu Gly 545 550 555 560 Gly Pro Ile Ser Asn Ala Leu Gln Lys Val Asp Val Gln Leu Ile Pro 565 570 575 Gln Asp Leu Cys Ser Glu Val Tyr Arg Tyr Gln Val Thr Pro Arg Met 580 585 590 Leu Cys Ala Gly Tyr Arg Lys Gly Lys Lys Asp Ala Cys Gln Gly Asp 595 600 605 Ser Gly Gly Pro Leu Val Cys Lys Ala Leu Ser Gly Arg Trp Phe Leu 610 615 620 Ala Gly Leu Val Ser Trp Gly Leu Gly Cys Gly Arg Pro Asn Tyr Phe 625 630 635 640 Gly Val Tyr Thr Arg Ile Thr Gly Val Ile Ser Trp Ile Gln Gln Val 645 650 655 Val Thr <210> SEQ ID NO 11 <211> LENGTH: 1656 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (268)...(1629) <223> OTHER INFORMATION: Nucleic acid encoding a transmembrane serine protease (MTSP-6) protein <400> SEQUENCE: 11 cgcccgggca ggtcagtaac actgtggcct actatctctt ccgtggtgcc atctacattt 60 ttgggactcg ggaattatga ctgtttttgg ttaatcgata ctgaatgcgc tttgtgtgga 120 ctgtcgaatt tcaaagattt accgtatgac caagatgcac ctgatgctac aagtataaat 180 aggggaacaa atgctttctg ttcttcctcg gctaaggagg tagaggtgga ggcggagccg 240 gatgtcagag gtcctgaaat agtcacc atg ggg gaa aat gat ccg cct gct gtt 294Met Gly Glu Asn Asp Pro Pro Ala Val 1 5 gaa gcc ccc ttc tca ttc cga tcg ctt ttt ggc ctt gat gat ttg aaa 342 Glu Ala Pro Phe Ser Phe Arg Ser Leu Phe Gly Leu Asp Asp Leu Lys 10 15 20 25 ata agt cct gtt gca cca gat gca gat gct gtt gct gca cag atc ctg 390Ile Ser Pro Val Ala Pro Asp Ala Asp Ala Val Ala Ala Gln Ile Leu 30 35 40 tca ctg ctg cca ttg aag ttt ttt cca atc atc gtc att ggg atc att 438 Ser Leu Leu Pro Leu Lys Phe Phe Pro Ile Ile Val Ile Gly Ile Ile 45 50 55 gca ttg ata tta gca ctg gcc att ggt ctg ggc atc cac ttc gac tgc 486Ala Leu Ile Leu Ala Leu Ala Ile Gly Leu Gly Ile His Phe Asp Cys 60 65 70 tca ggg aag tac aga tgt cgc tca tcc ttt aag tgt atc gag ctg ata 534 Ser Gly Lys Tyr Arg Cys Arg Ser Ser Phe Lys Cys Ile Glu Leu Ile 75 80 85 gct cga tgt gac gga gtc tcg gat tgc aaa gac ggg gag gac gag tac 582 Ala Arg Cys Asp Gly Val Ser Asp Cys Lys Asp Gly Glu Asp Glu Tyr 90 95 100 105 cgc tgt gtc cgg gtg ggt ggt cag aat gcc gtg ctc cag gtg ttc aca 630 Arg Cys Val Arg Val Gly Gly Gln Asn Ala Val Leu Gln Val Phe Thr 110 115 120 gct gct tcg tgg aag acc atg tgc tcc gat gac tgg aag ggt cac tac 678 Ala Ala Ser Trp Lys Thr Met Cys Ser Asp Asp Trp Lys Gly His Tyr 125 130 135 gca aat gtt gcc tgt gcc caa ctg ggt ttc cca agc tat gta agt tca 726 Ala Asn Val Ala Cys Ala Gln Leu Gly Phe Pro Ser Tyr Val Ser Ser 140 145 150 gat aac ctc aga gtg agc tcg cta gag ggg cag ttc cgg gag gag ttt 774 Asp Asn Leu Arg Val Ser Ser Leu Glu Gly Gln Phe Arg Glu Glu Phe 155 160 165 gtg tcc atc gat cac ctc ttg cca gat gac aag gtg act gca tta cac 822 Val Ser Ile Asp His Leu Leu Pro Asp Asp Lys Val Thr Ala Leu His 170 175 180 185 cac tca gta tat gtg agg gag gga tgt gcc tct ggc cac gtg gtt acc 870 His Ser Val Tyr Val Arg Glu Gly Cys Ala Ser Gly His Val Val Thr 190 195 200 ttg cag tgc aca gcc tgt ggt cat aga agg ggc tac agc tca cgc atc 918 Leu Gln Cys Thr Ala Cys Gly His Arg Arg Gly Tyr Ser Ser Arg Ile 205 210 215 gtg ggt gga aac atg tcc ttg ctc tcg cag tgg ccc tgg cag gcc agc 966 Val Gly Gly Asn Met Ser Leu Leu Ser Gln Trp Pro Trp Gln Ala Ser 220 225 230 ctt cag ttc cag ggc tac cac ctg tgc ggg ggc tct gtc atc acg ccc 1014 Leu Gln Phe Gln Gly Tyr His Leu Cys Gly Gly Ser Val Ile Thr Pro 235 240 245 ctg tgg atc atc act gct gca cac tgt gtt tat gac ttg tac ctc ccc 1062 Leu Trp Ile Ile Thr Ala Ala His Cys Val Tyr Asp Leu Tyr Leu Pro 250 255 260 265 aag tca tgg acc atc cag gtg ggt cta gtt tcc ctg ttg gac aat cca 1110 Lys Ser Trp Thr Ile Gln Val Gly Leu Val Ser Leu Leu Asp Asn Pro 270 275 280 gcc cca tcc cac ttg gtg gag aag att gtc tac cac agc aag tac aag 1158 Ala Pro Ser His Leu Val Glu Lys Ile Val Tyr His Ser Lys Tyr Lys 285 290 295 cca aag agg ctg ggc aat gac atc gcc ctt atg aag ctg gcc ggg cca 1206 Pro Lys Arg Leu Gly Asn Asp Ile Ala Leu Met Lys Leu Ala Gly Pro 300 305 310 ctc acg ttc aat gaa atg atc cag cct gtg tgc ctg ccc aac tct gaa 1254 Leu Thr Phe Asn Glu Met Ile Gln Pro Val Cys Leu Pro Asn Ser Glu 315 320 325 gag aac ttc ccc gat gga aaa gtg tgc tgg acg tca gga tgg ggg gcc 1302 Glu Asn Phe Pro Asp Gly Lys Val Cys Trp Thr Ser Gly Trp Gly Ala 330 335 340 345 aca gag gat gga ggt gac gcc tcc cct gtc ctg aac cac gcg gcc gtc 1350 Thr Glu Asp Gly Gly Asp Ala Ser Pro Val Leu Asn His Ala Ala Val 350 355 360 cct ttg att tcc aac aag atc tgc aac cac agg gac gtg tac ggt ggc 1398 Pro Leu Ile Ser Asn Lys Ile Cys Asn His Arg Asp Val Tyr Gly Gly 365 370 375 atc atc tcc ccc tcc atg ctc tgc gcg ggc tac ctg acg ggt ggc gtg 1446 Ile Ile Ser Pro Ser Met Leu Cys Ala Gly Tyr Leu Thr Gly Gly Val 380 385 390 gac agc tgc cag ggg gac agc ggg ggg ccc ctg gtg tgt caa gag agg 1494 Asp Ser Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Cys Gln Glu Arg 395 400 405 agg ctg tgg aag tta gtg gga gcg acc agc ttt ggc atc ggc tgc gca 1542 Arg Leu Trp Lys Leu Val Gly Ala Thr Ser Phe Gly Ile Gly Cys Ala 410 415 420 425 gag gtg aac aag cct ggg gtg tac acc cgt gtc acc tcc ttc ctg gac 1590 Glu Val Asn Lys Pro Gly Val Tyr Thr Arg Val Thr Ser Phe Leu Asp 430 435 440 tgg atc cac gag cag atg gag aga gac cta aaa acc tga agaggaaggg 1639 Trp Ile His Glu Gln Met Glu Arg Asp Leu Lys Thr * 445 450 gataagtagc cacctga 1656 <210> SEQ ID NO 12 <211> LENGTH: 453 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 12 Met Gly Glu Asn Asp Pro Pro Ala Val Glu Ala Pro Phe Ser Phe Arg 1 5 10 15 Ser Leu Phe Gly Leu Asp Asp Leu Lys Ile Ser Pro Val Ala Pro Asp 20 25 30 Ala Asp Ala Val Ala Ala Gln Ile Leu Ser Leu Leu Pro Leu Lys Phe 35 40 45 Phe Pro Ile Ile Val Ile Gly Ile Ile Ala Leu Ile Leu Ala Leu Ala 50 55 60 Ile Gly Leu Gly Ile His Phe Asp Cys Ser Gly Lys Tyr Arg Cys Arg 65 70 75 80 Ser Ser Phe Lys Cys Ile Glu Leu Ile Ala Arg Cys Asp Gly Val Ser 85 90 95 Asp Cys Lys Asp Gly Glu Asp Glu Tyr Arg Cys Val Arg Val Gly Gly 100 105 110 Gln Asn Ala Val Leu Gln Val Phe Thr Ala Ala Ser Trp Lys Thr Met 115 120 125 Cys Ser Asp Asp Trp Lys Gly His Tyr Ala Asn Val Ala Cys Ala Gln 130 135 140 Leu Gly Phe Pro Ser Tyr Val Ser Ser Asp Asn Leu Arg Val Ser Ser 145 150 155 160 Leu Glu Gly Gln Phe Arg Glu Glu Phe Val Ser Ile Asp His Leu Leu 165 170 175 Pro Asp Asp Lys Val Thr Ala Leu His His Ser Val Tyr Val Arg Glu 180 185 190 Gly Cys Ala Ser Gly His Val Val Thr Leu Gln Cys Thr Ala Cys Gly 195 200 205 His Arg Arg Gly Tyr Ser Ser Arg Ile Val Gly Gly Asn Met Ser Leu 210 215 220 Leu Ser Gln Trp Pro Trp Gln Ala Ser Leu Gln Phe Gln Gly Tyr His 225 230 235 240 Leu Cys Gly Gly Ser Val Ile Thr Pro Leu Trp Ile Ile Thr Ala Ala 245 250 255 His Cys Val Tyr Asp Leu Tyr Leu Pro Lys Ser Trp Thr Ile Gln Val 260 265 270 Gly Leu Val Ser Leu Leu Asp Asn Pro Ala Pro Ser His Leu Val Glu 275 280 285 Lys Ile Val Tyr His Ser Lys Tyr Lys Pro Lys Arg Leu Gly Asn Asp 290 295 300 Ile Ala Leu Met Lys Leu Ala Gly Pro Leu Thr Phe Asn Glu Met Ile 305 310 315 320 Gln Pro Val Cys Leu Pro Asn Ser Glu Glu Asn Phe Pro Asp Gly Lys 325 330 335 Val Cys Trp Thr Ser Gly Trp Gly Ala Thr Glu Asp Gly Gly Asp Ala 340 345 350 Ser Pro Val Leu Asn His Ala Ala Val Pro Leu Ile Ser Asn Lys Ile 355 360 365 Cys Asn His Arg Asp Val Tyr Gly Gly Ile Ile Ser Pro Ser Met Leu 370 375 380 Cys Ala Gly Tyr Leu Thr Gly Gly Val Asp Ser Cys Gln Gly Asp Ser 385 390 395 400 Gly Gly Pro Leu Val Cys Gln Glu Arg Arg Leu Trp Lys Leu Val Gly 405 410 415 Ala Thr Ser Phe Gly Ile Gly Cys Ala Glu Val Asn Lys Pro Gly Val 420 425 430 Tyr Thr Arg Val Thr Ser Phe Leu Asp Trp Ile His Glu Gln Met Glu 435 440 445 Arg Asp Leu Lys Thr 450 <210> SEQ ID NO 13 <211> LENGTH: 2100 <212> TYPE: DNA <213> ORGANISM: Homo sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (45)...(1361) <223> OTHER INFORMATION: Nucleic acid encoding MTSP7 <400> SEQUENCE: 13 agatcagatg gcgactgaat agaagctgcc ccagtcctgg gttc atg atg tac aca 56 Met Met Tyr Thr 1 cct gtt gaa ttt tca gaa gct gaa ttc tca cga gct gaa tat caa aga 104 Pro Val Glu Phe Ser Glu Ala Glu Phe Ser Arg Ala Glu Tyr Gln Arg 5 10 15 20 aag cag caa ttt tgg gac tca gta cgg cta gct ctt ttc aca tta gca 152 Lys Gln Gln Phe Trp Asp Ser Val Arg Leu Ala Leu Phe Thr Leu Ala 25 30 35 att gta gca atc ata gga att gca att ggt att gtt act cat ttt gtt 200 Ile Val Ala Ile Ile Gly Ile Ala Ile Gly Ile Val Thr His Phe Val 40 45 50 gtt gag gat gat aag tct ttc tat tac ctt gcc tct ttt aaa gtc aca 248 Val Glu Asp Asp Lys Ser Phe Tyr Tyr Leu Ala Ser Phe Lys Val Thr 55 60 65 aat atc aaa tat aaa gaa aat tat ggc ata aga tct tca aga gag ttt 296 Asn Ile Lys Tyr Lys Glu Asn Tyr Gly Ile Arg Ser Ser Arg Glu Phe 70 75 80 ata gaa agg agt cat cag att gaa aga atg atg tct agg ata ttt cga 344Ile Glu Arg Ser His Gln Ile Glu Arg Met Met Ser Arg Ile Phe Arg 85 90 95 100 cat tct tct gta ggc ggt cga ttt atc aaa tct cat gtt atc aaa tta 392 His Ser Ser Val Gly Gly Arg Phe Ile Lys Ser His Val Ile Lys Leu 105 110 115 agt cca gat gaa caa ggt gtg gat att ctt ata gtg ctc ata ttt cga 440Ser Pro Asp Glu Gln Gly Val Asp Ile Leu Ile Val Leu Ile Phe Arg 120 125 130 tac cca tct act gat agt gct gaa caa atc aag aaa aaa att gaa aag 488 Tyr Pro Ser Thr Asp Ser Ala Glu Gln Ile Lys Lys Lys Ile Glu Lys 135 140 145 gct tta tat caa agt ttg aag acc aaa caa ttg tct ttg acc ata aac 536Ala Leu Tyr Gln Ser Leu Lys Thr Lys Gln Leu Ser Leu Thr Ile Asn 150 155 160 aaa cca tca ttt aga ctc aca cct att gac agc aaa aag atg agg aat 584 Lys Pro Ser Phe Arg Leu Thr Pro Ile Asp Ser Lys Lys Met Arg Asn 165 170 175 180 ctt ctc aac agt cgc tgt gga ata agg atg aca tct tca aac atg cca 632 Leu Leu Asn Ser Arg Cys Gly Ile Arg Met Thr Ser Ser Asn Met Pro 185 190 195 tta cca gca tcc tct tct act caa aga att gtc caa gga agg gaa aca 680 Leu Pro Ala Ser Ser Ser Thr Gln Arg Ile Val Gln Gly Arg Glu Thr 200 205 210 gct atg gaa ggg gaa tgg cca tgg cag gcc agc ctc cag ctc ata ggg 728 Ala Met Glu Gly Glu Trp Pro Trp Gln Ala Ser Leu Gln Leu Ile Gly 215 220 225 tca ggc cat cag tgt gga gcc agc ctc atc agt aac aca tgg ctg ctc 776 Ser Gly His Gln Cys Gly Ala Ser Leu Ile Ser Asn Thr Trp Leu Leu 230 235 240 aca gca gct cac tgc ttt tgg aaa aat aaa gac cca act caa tgg att 824 Thr Ala Ala His Cys Phe Trp Lys Asn Lys Asp Pro Thr Gln Trp Ile 245 250 255 260 gct act ttt ggt gca act ata aca cca ccc gca gtg aaa cga aat gtg 872 Ala Thr Phe Gly Ala Thr Ile Thr Pro Pro Ala Val Lys Arg Asn Val 265 270 275 agg aaa att att ctt cat gag aat tac cat aga gaa aca aat gaa aat 920 Arg Lys Ile Ile Leu His Glu Asn Tyr His Arg Glu Thr Asn Glu Asn 280 285 290 gac att gct ttg gtt cag ctc tct act gga gtt gag ttt tca aat ata 968 Asp Ile Ala Leu Val Gln Leu Ser Thr Gly Val Glu Phe Ser Asn Ile 295 300 305 gtc cag aga gtt tgc ctc cca gac tca tct ata aag ttg cca cct aaa 1016 Val Gln Arg Val Cys Leu Pro Asp Ser Ser Ile Lys Leu Pro Pro Lys 310 315 320 aca agt gtg ttc gtc aca gga ttt gga tcc att gta gat gat gga cct 1064 Thr Ser Val Phe Val Thr Gly Phe Gly Ser Ile Val Asp Asp Gly Pro 325 330 335 340 ata caa aat aca ctt cgg caa gcc aga gtg gaa acc ata agc act gat 1112 Ile Gln Asn Thr Leu Arg Gln Ala Arg Val Glu Thr Ile Ser Thr Asp 345 350 355 gtg tgt aac aga aag gat gtg tat gat ggc ctg ata act cca gga atg 1160 Val Cys Asn Arg Lys Asp Val Tyr Asp Gly Leu Ile Thr Pro Gly Met 360 365 370 tta tgt gct gga ttc atg gaa gga aaa ata gat gca tgt aag gga gat 1208 Leu Cys Ala Gly Phe Met Glu Gly Lys Ile Asp Ala Cys Lys Gly Asp 375 380 385 tct ggt gga cct ctg gtt tat gat aat cat gac atc tgg tac att gta 1256 Ser Gly Gly Pro Leu Val Tyr Asp Asn His Asp Ile Trp Tyr Ile Val 390 395 400 ggt ata gta agt tgg gga caa tca tgt gca ctt ccc aaa aaa cct gga 1304 Gly Ile Val Ser Trp Gly Gln Ser Cys Ala Leu Pro Lys Lys Pro Gly 405 410 415 420 gtc tac acc aga gta act aag tat cga gat tgg att gcc tca aag act 1352 Val Tyr Thr Arg Val Thr Lys Tyr Arg Asp Trp Ile Ala Ser Lys Thr 425 430 435 ggt atg tag tgtggattgt ccatgagtta tacacatggc acacagagct 1401 Gly Met * gatactcctg cgtattttgt attgtttaaa ttcatttact ttggattagt gcttttgcta 1461 gatgtcaaga agcccttcag acccagacaa atctaatatc ctgaggtggc ctttacatac 1521 gtaggaccaa accctctcta ccatgaggga agaagacaca gcaaatgaca gacagcacct 1581 attccttact cacaagggaa actgcttgtg atacttccta ataagataaa taagtggttt 1641 ccctcaattg aagacaggaa catcattttc cacaggatat gaagagctgc cagtaatgcc 1701 aaaatcttac ctcatataat acctggagca tgtgagattc ttctagtgaa aaagaacagt 1761 cttccctgaa gactcagggc ttcaacattc tagaactgat aagtggacct tcagtgtgca 1821 agaatggaga agcatgggat ttgcattatg acttgaactg ggcttatatc taataataca 1881 gagcactatc actaacctca acagttgaca ttttaaaagt ttttaaatgt atctgaactt 1941 gctgttaaca cagtgttata actcaagcac tagcttcagg aagcatgttg tgttgttaag 2001 aagcttttct gatttattct ttaacagcat cttgccatct atatgttagt agcagttggc 2061 ccagaaagga caaaaaaaaa aaaaaaaaaa aaaaaaaaa 2100 <210> SEQ ID NO 14 <211> LENGTH: 438 <212> TYPE: PRT <213> ORGANISM: Homo sapien <400> SEQUENCE: 14 Met Met Tyr Thr Pro Val Glu Phe Ser Glu Ala Glu Phe Ser Arg Ala 1 5 10 15 Glu Tyr Gln Arg Lys Gln Gln Phe Trp Asp Ser Val Arg Leu Ala Leu 20 25 30 Phe Thr Leu Ala Ile Val Ala Ile Ile Gly Ile Ala Ile Gly Ile Val 35 40 45 Thr His Phe Val Val Glu Asp Asp Lys Ser Phe Tyr Tyr Leu Ala Ser 50 55 60 Phe Lys Val Thr Asn Ile Lys Tyr Lys Glu Asn Tyr Gly Ile Arg Ser 65 70 75 80 Ser Arg Glu Phe Ile Glu Arg Ser His Gln Ile Glu Arg Met Met Ser 85 90 95 Arg Ile Phe Arg His Ser Ser Val Gly Gly Arg Phe Ile Lys Ser His 100 105 110 Val Ile Lys Leu Ser Pro Asp Glu Gln Gly Val Asp Ile Leu Ile Val 115 120 125 Leu Ile Phe Arg Tyr Pro Ser Thr Asp Ser Ala Glu Gln Ile Lys Lys 130 135 140 Lys Ile Glu Lys Ala Leu Tyr Gln Ser Leu Lys Thr Lys Gln Leu Ser 145 150 155 160 Leu Thr Ile Asn Lys Pro Ser Phe Arg Leu Thr Pro Ile Asp Ser Lys 165 170 175 Lys Met Arg Asn Leu Leu Asn Ser Arg Cys Gly Ile Arg Met Thr Ser 180 185 190 Ser Asn Met Pro Leu Pro Ala Ser Ser Ser Thr Gln Arg Ile Val Gln 195 200 205 Gly Arg Glu Thr Ala Met Glu Gly Glu Trp Pro Trp Gln Ala Ser Leu 210 215 220 Gln Leu Ile Gly Ser Gly His Gln Cys Gly Ala Ser Leu Ile Ser Asn 225 230 235 240 Thr Trp Leu Leu Thr Ala Ala His Cys Phe Trp Lys Asn Lys Asp Pro 245 250 255 Thr Gln Trp Ile Ala Thr Phe Gly Ala Thr Ile Thr Pro Pro Ala Val 260 265 270 Lys Arg Asn Val Arg Lys Ile Ile Leu His Glu Asn Tyr His Arg Glu 275 280 285 Thr Asn Glu Asn Asp Ile Ala Leu Val Gln Leu Ser Thr Gly Val Glu 290 295 300 Phe Ser Asn Ile Val Gln Arg Val Cys Leu Pro Asp Ser Ser Ile Lys 305 310 315 320 Leu Pro Pro Lys Thr Ser Val Phe Val Thr Gly Phe Gly Ser Ile Val 325 330 335 Asp Asp Gly Pro Ile Gln Asn Thr Leu Arg Gln Ala Arg Val Glu Thr 340 345 350 Ile Ser Thr Asp Val Cys Asn Arg Lys Asp Val Tyr Asp Gly Leu Ile 355 360 365 Thr Pro Gly Met Leu Cys Ala Gly Phe Met Glu Gly Lys Ile Asp Ala 370 375 380 Cys Lys Gly Asp Ser Gly Gly Pro Leu Val Tyr Asp Asn His Asp Ile 385 390 395 400 Trp Tyr Ile Val Gly Ile Val Ser Trp Gly Gln Ser Cys Ala Leu Pro 405 410 415 Lys Lys Pro Gly Val Tyr Thr Arg Val Thr Lys Tyr Arg Asp Trp Ile 420 425 430 Ala Ser Lys Thr Gly Met 435 <210> SEQ ID NO 15 <211> LENGTH: 702 <212> TYPE: DNA <213> ORGANISM: Homo sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (1)...(702) <223> OTHER INFORMATION: Nucleotide sequence encoding MTSP-7 Protease Domain <400> SEQUENCE: 15 att gtc caa gga agg gaa aca gct atg gaa ggg gaa tgg cca tgg cag 48 Ile Val Gln Gly Arg Glu Thr Ala Met Glu Gly Glu Trp Pro Trp Gln 1 5 10 15 gcc agc ctc cag ctc ata ggg tca ggc cat cag tgt gga gcc agc ctc 96 Ala Ser Leu Gln Leu Ile Gly Ser Gly His Gln Cys Gly Ala Ser Leu 20 25 30 atc agt aac aca tgg ctg ctc aca gca gct cac tgc ttt tgg aaa aat 144 Ile Ser Asn Thr Trp Leu Leu Thr Ala Ala His Cys Phe Trp Lys Asn 35 40 45 aaa gac cca act caa tgg att gct act ttt ggt gca act ata aca cca 192 Lys Asp Pro Thr Gln Trp Ile Ala Thr Phe Gly Ala Thr Ile Thr Pro 50 55 60 ccc gca gtg aaa cga aat gtg agg aaa att att ctt cat gag aat tac 240 Pro Ala Val Lys Arg Asn Val Arg Lys Ile Ile Leu His Glu Asn Tyr 65 70 75 80 cat aga gaa aca aat gaa aat gac att gct ttg gtt cag ctc tct act 288His Arg Glu Thr Asn Glu Asn Asp Ile Ala Leu Val Gln Leu Ser Thr 85 90 95 gga gtt gag ttt tca aat ata gtc cag aga gtt tgc ctc cca gac tca 336Gly Val Glu Phe Ser Asn Ile Val Gln Arg Val Cys Leu Pro Asp Ser 100 105 110 tct ata aag ttg cca cct aaa aca agt gtg ttc gtc aca gga ttt gga 384Ser Ile Lys Leu Pro Pro Lys Thr Ser Val Phe Val Thr Gly Phe Gly 115 120 125 tcc att gta gat gat gga cct ata caa aat aca ctt cgg caa gcc aga 432 Ser Ile Val Asp Asp Gly Pro Ile Gln Asn Thr Leu Arg Gln Ala Arg 130 135 140 gtg gaa acc ata agc act gat gtg tgt aac aga aag gat gtg tat gat 480 Val Glu Thr Ile Ser Thr Asp Val Cys Asn Arg Lys Asp Val Tyr Asp 145 150 155 160 ggc ctg ata act cca gga atg tta tgt gct gga ttc atg gaa gga aaa 528 Gly Leu Ile Thr Pro Gly Met Leu Cys Ala Gly Phe Met Glu Gly Lys 165 170 175 ata gat gca tgt aag gga gat tct ggt gga cct ctg gtt tat gat aat 576 Ile Asp Ala Cys Lys Gly Asp Ser Gly Gly Pro Leu Val Tyr Asp Asn 180 185 190 cat gac atc tgg tac att gta ggt ata gta agt tgg gga caa tca tgt 624 His Asp Ile Trp Tyr Ile Val Gly Ile Val Ser Trp Gly Gln Ser Cys 195 200 205 gca ctt ccc aaa aaa cct gga gtc tac acc aga gta act aag tat cga 672 Ala Leu Pro Lys Lys Pro Gly Val Tyr Thr Arg Val Thr Lys Tyr Arg 210 215 220 gat tgg att gcc tca aag act ggt atg tag 702 Asp Trp Ile Ala Ser Lys Thr Gly Met * 225 230 <210> SEQ ID NO 16 <211> LENGTH: 233 <212> TYPE: PRT <213> ORGANISM: Homo sapien <400> SEQUENCE: 16 Ile Val Gln Gly Arg Glu Thr Ala Met Glu Gly Glu Trp Pro Trp Gln 1 5 10 15 Ala Ser Leu Gln Leu Ile Gly Ser Gly His Gln Cys Gly Ala Ser Leu 20 25 30 Ile Ser Asn Thr Trp Leu Leu Thr Ala Ala His Cys Phe Trp Lys Asn 35 40 45 Lys Asp Pro Thr Gln Trp Ile Ala Thr Phe Gly Ala Thr Ile Thr Pro 50 55 60 Pro Ala Val Lys Arg Asn Val Arg Lys Ile Ile Leu His Glu Asn Tyr 65 70 75 80 His Arg Glu Thr Asn Glu Asn Asp Ile Ala Leu Val Gln Leu Ser Thr 85 90 95 Gly Val Glu Phe Ser Asn Ile Val Gln Arg Val Cys Leu Pro Asp Ser 100 105 110 Ser Ile Lys Leu Pro Pro Lys Thr Ser Val Phe Val Thr Gly Phe Gly 115 120 125 Ser Ile Val Asp Asp Gly Pro Ile Gln Asn Thr Leu Arg Gln Ala Arg 130 135 140 Val Glu Thr Ile Ser Thr Asp Val Cys Asn Arg Lys Asp Val Tyr Asp 145 150 155 160 Gly Leu Ile Thr Pro Gly Met Leu Cys Ala Gly Phe Met Glu Gly Lys 165 170 175 Ile Asp Ala Cys Lys Gly Asp Ser Gly Gly Pro Leu Val Tyr Asp Asn 180 185 190 His Asp Ile Trp Tyr Ile Val Gly Ile Val Ser Trp Gly Gln Ser Cys 195 200 205 Ala Leu Pro Lys Lys Pro Gly Val Tyr Thr Arg Val Thr Lys Tyr Arg 210 215 220 Asp Trp Ile Ala Ser Lys Thr Gly Met 225 230 <210> SEQ ID NO 17 <211> LENGTH: 777 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (1)...(729) <223> OTHER INFORMATION: Nucleotide sequence encoding MTSP9, including protease domain (31-729) <400> SEQUENCE: 17 aaa cga gtt gtt cca tta aac gtc aac aga ata gca tct gga gtc att 48 Lys Arg Val Val Pro Leu Asn Val Asn Arg Ile Ala Ser Gly Val Ile 1 5 10 15 gca ccc aag gcg gcc tgg cct tgg caa gct tcc ctt cag tat gat aac 96 Ala Pro Lys Ala Ala Trp Pro Trp Gln Ala Ser Leu Gln Tyr Asp Asn 20 25 30 atc cat cag tgt ggg gcc acc ttg att agt aac aca tgg ctt gtc act 144 Ile His Gln Cys Gly Ala Thr Leu Ile Ser Asn Thr Trp Leu Val Thr 35 40 45 gca gca cac tgc ttc cag aag tat aaa aat cca cat caa tgg act gtt 192 Ala Ala His Cys Phe Gln Lys Tyr Lys Asn Pro His Gln Trp Thr Val 50 55 60 agt ttt gga aca aaa atc aac cct ccc tta atg aaa aga aat gtc aga 240 Ser Phe Gly Thr Lys Ile Asn Pro Pro Leu Met Lys Arg Asn Val Arg 65 70 75 80 aga ttt att atc cat gag aag tac cgc tct gca gca aga gag tac gac 288 Arg Phe Ile Ile His Glu Lys Tyr Arg Ser Ala Ala Arg Glu Tyr Asp 85 90 95 att gct gtt gtg cag gtc tct tcc aga gtc acc ttt tcg gat gac ata 336 Ile Ala Val Val Gln Val Ser Ser Arg Val Thr Phe Ser Asp Asp Ile 100 105 110 cgc cgg att tgt ttg cca gaa gcc tct gca tcc ttc caa cca aat ttg 384Arg Arg Ile Cys Leu Pro Glu Ala Ser Ala Ser Phe Gln Pro Asn Leu 115 120 125 act gtc cac atc aca gga ttt gga gca ctt tac tat ggt ggg gaa tcc 432Thr Val His Ile Thr Gly Phe Gly Ala Leu Tyr Tyr Gly Gly Glu Ser 130 135 140 caa aat gat ctc cga gaa gcc aga gtg aaa atc ata agt gac gat gtc 480Gln Asn Asp Leu Arg Glu Ala Arg Val Lys Ile Ile Ser Asp Asp Val 145 150 155 160 tgc aag caa cca cag gtg tat ggc aat gat ata aaa cct gga atg ttc 528 Cys Lys Gln Pro Gln Val Tyr Gly Asn Asp Ile Lys Pro Gly Met Phe 165 170 175 tgt gcc gga tat atg gaa gga att tat gat gcc tgc agg ggt gat tct 576 Cys Ala Gly Tyr Met Glu Gly Ile Tyr Asp Ala Cys Arg Gly Asp Ser 180 185 190 ggg gga cct tta gtc aca agg gat ctg aaa gat acg tgg tat ctc att 624 Gly Gly Pro Leu Val Thr Arg Asp Leu Lys Asp Thr Trp Tyr Leu Ile 195 200 205 gga att gta agc tgg gga gat aac tgt ggt caa aag gac aag cct gga 672 Gly Ile Val Ser Trp Gly Asp Asn Cys Gly Gln Lys Asp Lys Pro Gly 210 215 220 gtc tac aca caa gtg act tat tac cga aac tgg att gct tca aaa aca 720 Val Tyr Thr Gln Val Thr Tyr Tyr Arg Asn Trp Ile Ala Ser Lys Thr 225 230 235 240 ggc atc taa ttcacgataa aagttaaaca aagaaagctg tatgcaggtc atatatgc 777 Gly Ile <210> SEQ ID NO 18 <211> LENGTH: 242 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: SITE <222> LOCATION: (11)...(242) <223> OTHER INFORMATION: MTSP9 protease domain <400> SEQUENCE: 18 Lys Arg Val Val Pro Leu Asn Val Asn Arg Ile Ala Ser Gly Val Ile 1 5 10 15 Ala Pro Lys Ala Ala Trp Pro Trp Gln Ala Ser Leu Gln Tyr Asp Asn 20 25 30 Ile His Gln Cys Gly Ala Thr Leu Ile Ser Asn Thr Trp Leu Val Thr 35 40 45 Ala Ala His Cys Phe Gln Lys Tyr Lys Asn Pro His Gln Trp Thr Val 50 55 60 Ser Phe Gly Thr Lys Ile Asn Pro Pro Leu Met Lys Arg Asn Val Arg 65 70 75 80 Arg Phe Ile Ile His Glu Lys Tyr Arg Ser Ala Ala Arg Glu Tyr Asp 85 90 95 Ile Ala Val Val Gln Val Ser Ser Arg Val Thr Phe Ser Asp Asp Ile 100 105 110 Arg Arg Ile Cys Leu Pro Glu Ala Ser Ala Ser Phe Gln Pro Asn Leu 115 120 125 Thr Val His Ile Thr Gly Phe Gly Ala Leu Tyr Tyr Gly Gly Glu Ser 130 135 140 Gln Asn Asp Leu Arg Glu Ala Arg Val Lys Ile Ile Ser Asp Asp Val 145 150 155 160 Cys Lys Gln Pro Gln Val Tyr Gly Asn Asp Ile Lys Pro Gly Met Phe 165 170 175 Cys Ala Gly Tyr Met Glu Gly Ile Tyr Asp Ala Cys Arg Gly Asp Ser 180 185 190 Gly Gly Pro Leu Val Thr Arg Asp Leu Lys Asp Thr Trp Tyr Leu Ile 195 200 205 Gly Ile Val Ser Trp Gly Asp Asn Cys Gly Gln Lys Asp Lys Pro Gly 210 215 220 Val Tyr Thr Gln Val Thr Tyr Tyr Arg Asn Trp Ile Ala Ser Lys Thr 225 230 235 240 Gly Ile <210> SEQ ID NO 19 <211> LENGTH: 3316 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (1)...(3282) <223> OTHER INFORMATION: Nucleotide sequence encoding MTSP12, including MTSP12-PD1, MTSP12-PD2, and MTSP12-PD3 protease domains <400> SEQUENCE: 19 atg gag ccc act gtg gct aac gta cac ctc gtg ccc agg aca acc aag 48 Met Glu Pro Thr Val Ala Asn Val His Leu Val Pro Arg Thr Thr Lys 1 5 10 15 gaa gtc ccc gct ctg gat gcc gcg tgc tgt cga gcg gcc acc att ggc 96 Glu Val Pro Ala Leu Asp Ala Ala Cys Cys Arg Ala Ala Thr Ile Gly 20 25 30 gtg gtg gcc acc agc ctt gtc gtc ctc acc ctg gga gtc ctt ttg gcc 144 Val Val Ala Thr Ser Leu Val Val Leu Thr Leu Gly Val Leu Leu Ala 35 40 45 ttc ctc tct aca cag ggc ttc cac gtg gac cac acg gcc gag ctg cgg 192 Phe Leu Ser Thr Gln Gly Phe His Val Asp His Thr Ala Glu Leu Arg 50 55 60 gga atc cgg tgg acc agc agt ttg cgg cgg gag acc tcg gac tat cac 240 Gly Ile Arg Trp Thr Ser Ser Leu Arg Arg Glu Thr Ser Asp Tyr His 65 70 75 80 cgc acg ctg acg ccc acc ctg gag gca ctg ttt gta agt agt ttt cag 288Arg Thr Leu Thr Pro Thr Leu Glu Ala Leu Phe Val Ser Ser Phe Gln 85 90 95 aag aca gag tta gag gca agc tgc gtg ggt tgc tcg gta ctg aat tat 336 Lys Thr Glu Leu Glu Ala Ser Cys Val Gly Cys Ser Val Leu Asn Tyr 100 105 110 agg gat ggg aac tcc agt gtc ctc gta cat ttc cag ctg cac ttt ctg 384Arg Asp Gly Asn Ser Ser Val Leu Val His Phe Gln Leu His Phe Leu 115 120 125 ctg cga ccc ctc cag acg ctg agc ctg ggc ctg gag gag gag cta ttg 432Leu Arg Pro Leu Gln Thr Leu Ser Leu Gly Leu Glu Glu Glu Leu Leu 130 135 140 cag cga ggg atc cgg gca agg ctg cgg gag cac ggc atc tcc ctg gct 480Gln Arg Gly Ile Arg Ala Arg Leu Arg Glu His Gly Ile Ser Leu Ala 145 150 155 160 gcc tat ggc aca att gtg tcg gct gag ctc aca ggg aga cat aag ggg 528Ala Tyr Gly Thr Ile Val Ser Ala Glu Leu Thr Gly Arg His Lys Gly 165 170 175 ccc ttg gca gaa aga gac ttc aaa tca ggc cgc tgt cca ggg aac tcc 576 Pro Leu Ala Glu Arg Asp Phe Lys Ser Gly Arg Cys Pro Gly Asn Ser 180 185 190 ttt tcc tgc ggg aac agc cag tgt gtg acc aag gtg aac ccg gag tgt 624 Phe Ser Cys Gly Asn Ser Gln Cys Val Thr Lys Val Asn Pro Glu Cys 195 200 205 gac gac cag gag gac tgc tcc gat ggg tcc gac gag gcg cac tgc gag 672 Asp Asp Gln Glu Asp Cys Ser Asp Gly Ser Asp Glu Ala His Cys Glu 210 215 220 tgt ggc ttg cag cct gcc tgg agg atg gcc ggc agg atc gtg ggc ggc 720 Cys Gly Leu Gln Pro Ala Trp Arg Met Ala Gly Arg Ile Val Gly Gly 225 230 235 240 atg gaa gca tcc ccg ggg gag ttt ccg tgg caa gcc agc ctt cga gag 768 Met Glu Ala Ser Pro Gly Glu Phe Pro Trp Gln Ala Ser Leu Arg Glu 245 250 255 aac aag gag cac ttc tgt ggg gcc gcc atc atc aac gcc agg tgg ctg 816 Asn Lys Glu His Phe Cys Gly Ala Ala Ile Ile Asn Ala Arg Trp Leu 260 265 270 gtg tct gct gct cac tgc ttc aat gag ttc caa gac ccg acg aag tgg 864 Val Ser Ala Ala His Cys Phe Asn Glu Phe Gln Asp Pro Thr Lys Trp 275 280 285 gtg gcc tac gtg ggt gcg acc tac ctc agc ggc tcg gag gcc agc acc 912 Val Ala Tyr Val Gly Ala Thr Tyr Leu Ser Gly Ser Glu Ala Ser Thr 290 295 300 gtg cgg gcc cag gtg gtc cag atc gtc aag cac ccc ctg tac aac gcg 960 Val Arg Ala Gln Val Val Gln Ile Val Lys His Pro Leu Tyr Asn Ala 305 310 315 320 gac acg gcc gac ttt gac gtg gct gtg ctg gag ctg acc agc cct ctg 1008 Asp Thr Ala Asp Phe Asp Val Ala Val Leu Glu Leu Thr Ser Pro Leu 325 330 335 cct ttc ggc cgg cac atc cag ccc gtg tgc ctc ccg gct gcc aca cac 1056 Pro Phe Gly Arg His Ile Gln Pro Val Cys Leu Pro Ala Ala Thr His 340 345 350 atc ttc cca ccc agc aag aag tgc ctg atc tca ggc tgg ggc tac ctc 1104 Ile Phe Pro Pro Ser Lys Lys Cys Leu Ile Ser Gly Trp Gly Tyr Leu 355 360 365 aag gag gac ttc ctg gtc aag cca ggg gtg ctg cag aaa gcc act gtg 1152 Lys Glu Asp Phe Leu Val Lys Pro Gly Val Leu Gln Lys Ala Thr Val 370 375 380 gag ctg ctg gac cag gca ctg tgt gcc agc ttg tac ggc cat tca ctc 1200 Glu Leu Leu Asp Gln Ala Leu Cys Ala Ser Leu Tyr Gly His Ser Leu 385 390 395 400 act gac agg atg gtg tgc gct ggc tac ctg gac ggg aag gtg gac tcc 1248 Thr Asp Arg Met Val Cys Ala Gly Tyr Leu Asp Gly Lys Val Asp Ser 405 410 415 tgc cag ggt gac tca gga gga ccc ctg gtc tgc gag gag ccc tct ggc 1296 Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Cys Glu Glu Pro Ser Gly 420 425 430 cgg ttc tct ctg gct ggc atc gtg agc tgg gga atc ggg tgt gcg gaa 1344 Arg Phe Ser Leu Ala Gly Ile Val Ser Trp Gly Ile Gly Cys Ala Glu 435 440 445 gcc cgg cgt cca ggg gtc tat gcc cga gtc acc agg cta cgt gac tgg 1392 Ala Arg Arg Pro Gly Val Tyr Ala Arg Val Thr Arg Leu Arg Asp Trp 450 455 460 atc ctg gag gcc acc acc aaa gcc agc atg cct ctg gcc ccc acc atg 1440 Ile Leu Glu Ala Thr Thr Lys Ala Ser Met Pro Leu Ala Pro Thr Met 465 470 475 480 gct cct gcc cct gcc gcc ccc agc aca gcc tgg ccc acc agt cct gag 1488 Ala Pro Ala Pro Ala Ala Pro Ser Thr Ala Trp Pro Thr Ser Pro Glu 485 490 495 agc cct gtt gtc agc acc ccc acc aaa tcg atg cag gcc ctc agt acc 1536 Ser Pro Val Val Ser Thr Pro Thr Lys Ser Met Gln Ala Leu Ser Thr 500 505 510 gtg cct ctt gac tgg gtc acc gtt cct aag cta caa gaa tgt ggg gcc 1584 Val Pro Leu Asp Trp Val Thr Val Pro Lys Leu Gln Glu Cys Gly Ala 515 520 525 agg cct gca atg gag aag ccc acc cgg gtc gtg ggc ggg ttc gga gct 1632 Arg Pro Ala Met Glu Lys Pro Thr Arg Val Val Gly Gly Phe Gly Ala 530 535 540 gcc tcc ggg gag gtg ccc tgg cag gtc agc ctg aag gaa ggg tcc cgg 1680 Ala Ser Gly Glu Val Pro Trp Gln Val Ser Leu Lys Glu Gly Ser Arg 545 550 555 560 cac ttc tgc gga gca act gtg gtg ggg gac cgc tgg ctg ctg tct gcc 1728 His Phe Cys Gly Ala Thr Val Val Gly Asp Arg Trp Leu Leu Ser Ala 565 570 575 gcc cac tgc ttc aac cac acg aag gtg gag cag gtt cgg gcc cac ctg 1776 Ala His Cys Phe Asn His Thr Lys Val Glu Gln Val Arg Ala His Leu 580 585 590 ggc act gcg tcc ctc ctg ggc ctg ggc ggg agc ccg gtg aag atc ggg 1824 Gly Thr Ala Ser Leu Leu Gly Leu Gly Gly Ser Pro Val Lys Ile Gly 595 600 605 ctg cgg cgg gta gtg ctg cac ccc ctc tac aac cct ggc atc ctg gac 1872 Leu Arg Arg Val Val Leu His Pro Leu Tyr Asn Pro Gly Ile Leu Asp 610 615 620 ttc gac ctg gct gtc ctg gag ctg gcc agc ccc ctg gcc ttc aac aaa 1920 Phe Asp Leu Ala Val Leu Glu Leu Ala Ser Pro Leu Ala Phe Asn Lys 625 630 635 640 tac atc cag cct gtc tgc ctg ccc ctg gcc atc cgg aag ttc cct gtg 1968 Tyr Ile Gln Pro Val Cys Leu Pro Leu Ala Ile Arg Lys Phe Pro Val 645 650 655 ggc cgg aag tgc atg atc tcc gga tgg gga aat acg cag gaa gga aat 2016 Gly Arg Lys Cys Met Ile Ser Gly Trp Gly Asn Thr Gln Glu Gly Asn 660 665 670 gcc acc aag ccc gag ctc ctg cag aag gcg tcc gtg ggc atc ata gac 2064 Ala Thr Lys Pro Glu Leu Leu Gln Lys Ala Ser Val Gly Ile Ile Asp 675 680 685 cag aaa acc tgt agt gtg ctc tac aac ttc tcc ctc aca gac cgc atg 2112 Gln Lys Thr Cys Ser Val Leu Tyr Asn Phe Ser Leu Thr Asp Arg Met 690 695 700 atc tgc gca ggc ttc ctg gaa ggc aaa gtc gac tcc tgc cag ggt gac 2160 Ile Cys Ala Gly Phe Leu Glu Gly Lys Val Asp Ser Cys Gln Gly Asp 705 710 715 720 tct ggg ggc ccc ctg gcc tgc gag gag gcc cct ggc gtg ttt tat ctg 2208 Ser Gly Gly Pro Leu Ala Cys Glu Glu Ala Pro Gly Val Phe Tyr Leu 725 730 735 gca ggg atc gtg agc tgg ggt att ggc tgc gct cag gtt aag aag ccg 2256 Ala Gly Ile Val Ser Trp Gly Ile Gly Cys Ala Gln Val Lys Lys Pro 740 745 750 ggc gtg tac acg cgc atc acc agg cta aag ggc tgg atc ctg gag atc 2304 Gly Val Tyr Thr Arg Ile Thr Arg Leu Lys Gly Trp Ile Leu Glu Ile 755 760 765 atg tcc tcc cag ccc ctt ccc atg tct ccc ccc tcg acc aca agg atg 2352 Met Ser Ser Gln Pro Leu Pro Met Ser Pro Pro Ser Thr Thr Arg Met 770 775 780 ctg gcc acc acc agc ccc agg acg aca gct ggc ctc aca gtc ccg ggg 2400 Leu Ala Thr Thr Ser Pro Arg Thr Thr Ala Gly Leu Thr Val Pro Gly 785 790 795 800 gcc aca ccc agc aga ccc acc cct ggg gct gcc agc agg gtg acg ggc 2448 Ala Thr Pro Ser Arg Pro Thr Pro Gly Ala Ala Ser Arg Val Thr Gly 805 810 815 caa cct gcc aac tca acc tta tct gcc gtg agc acc act gct agg gga 2496 Gln Pro Ala Asn Ser Thr Leu Ser Ala Val Ser Thr Thr Ala Arg Gly 820 825 830 cag acg cca ttt cca gac gcc ccg gag gcc acc aca cac acc cag cta 2544 Gln Thr Pro Phe Pro Asp Ala Pro Glu Ala Thr Thr His Thr Gln Leu 835 840 845 cca gac tgt ggc ctg gcg ccg gcc gcg ctc acc agg att gtg ggc ggc 2592 Pro Asp Cys Gly Leu Ala Pro Ala Ala Leu Thr Arg Ile Val Gly Gly 850 855 860 agc gca gcg ggc cgt ggg gag tgg ccg tgg cag gtg ggc ctg tgg ctg 2640 Ser Ala Ala Gly Arg Gly Glu Trp Pro Trp Gln Val Gly Leu Trp Leu 865 870 875 880 cgg cgc cgg gaa cac cgt tgc ggg gcc gtg ctg gtg gca gag agg tgg 2688 Arg Arg Arg Glu His Arg Cys Gly Ala Val Leu Val Ala Glu Arg Trp 885 890 895 ctg ctg tcg gcg gcg cac tgc ttc gac gtc tac ggg gac ccc aag cag 2736 Leu Leu Ser Ala Ala His Cys Phe Asp Val Tyr Gly Asp Pro Lys Gln 900 905 910 tgg gcg gcc ttc cta ggc acg ccg ttc ctg agc ggc gcg gag ggg cag 2784 Trp Ala Ala Phe Leu Gly Thr Pro Phe Leu Ser Gly Ala Glu Gly Gln 915 920 925 ctg gag cgc gtg gcg cgc atc tac aag cac ccg ttc tac aat ctc tac 2832 Leu Glu Arg Val Ala Arg Ile Tyr Lys His Pro Phe Tyr Asn Leu Tyr 930 935 940 acg ctc gac tac gac gtg gcg ctt ctg gag ctg gcg ggg ccg gtg cgt 2880 Thr Leu Asp Tyr Asp Val Ala Leu Leu Glu Leu Ala Gly Pro Val Arg 945 950 955 960 cgc agc cgc ctg gtg cgt ccc atc tgc ctg ccc gag ccc gcg ccg cga 2928 Arg Ser Arg Leu Val Arg Pro Ile Cys Leu Pro Glu Pro Ala Pro Arg 965 970 975 ccc ccg gac ggc acg cgc tgc gtc atc acc ggc tgg ggc tcg gtg cgc 2976 Pro Pro Asp Gly Thr Arg Cys Val Ile Thr Gly Trp Gly Ser Val Arg 980 985 990 gaa gga ggc tcc atg gcg cgg cag ctg cag aag gcg gcc gtg cgc ctc 3024 Glu Gly Gly Ser Met Ala Arg Gln Leu Gln Lys Ala Ala Val Arg Leu 995 1000 1005 ctc agc gag cag acc tgc cgc cgc ttc tac cca gtg cag atc agc agc 3072 Leu Ser Glu Gln Thr Cys Arg Arg Phe Tyr Pro Val Gln Ile Ser Ser 1010 1015 1020 cgc atg ctg tgt gcc ggc ttc ccg cag ggt ggc gtg gac agc tgc tcg 3120 Arg Met Leu Cys Ala Gly Phe Pro Gln Gly Gly Val Asp Ser Cys Ser 1025 1030 1035 1040 ggt gac gct ggg gga ccc ctg gcc tgc agg gag ccc tct gga cgg tgg 3168 Gly Asp Ala Gly Gly Pro Leu Ala Cys Arg Glu Pro Ser Gly Arg Trp 1045 1050 1055 gtg cta act ggg gtc act agc tgg ggc tat ggc tgt ggc cgg ccc cac 3216 Val Leu Thr Gly Val Thr Ser Trp Gly Tyr Gly Cys Gly Arg Pro His 1060 1065 1070 ttc cca ggt gtc tat acc cgg gtg gca gct gtg aga ggc tgg ata gga 3264 Phe Pro Gly Val Tyr Thr Arg Val Ala Ala Val Arg Gly Trp Ile Gly 1075 1080 1085 cag cac atc cag gag tga ccaccacgtg actgcccagg ccgagactct 3312 Gln His Ile Gln Glu * 1090 acgt 3316 <210> SEQ ID NO 20 <211> LENGTH: 1093 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 20 Met Glu Pro Thr Val Ala Asn Val His Leu Val Pro Arg Thr Thr Lys 1 5 10 15 Glu Val Pro Ala Leu Asp Ala Ala Cys Cys Arg Ala Ala Thr Ile Gly 20 25 30 Val Val Ala Thr Ser Leu Val Val Leu Thr Leu Gly Val Leu Leu Ala 35 40 45 Phe Leu Ser Thr Gln Gly Phe His Val Asp His Thr Ala Glu Leu Arg 50 55 60 Gly Ile Arg Trp Thr Ser Ser Leu Arg Arg Glu Thr Ser Asp Tyr His 65 70 75 80 Arg Thr Leu Thr Pro Thr Leu Glu Ala Leu Phe Val Ser Ser Phe Gln 85 90 95 Lys Thr Glu Leu Glu Ala Ser Cys Val Gly Cys Ser Val Leu Asn Tyr 100 105 110 Arg Asp Gly Asn Ser Ser Val Leu Val His Phe Gln Leu His Phe Leu 115 120 125 Leu Arg Pro Leu Gln Thr Leu Ser Leu Gly Leu Glu Glu Glu Leu Leu 130 135 140 Gln Arg Gly Ile Arg Ala Arg Leu Arg Glu His Gly Ile Ser Leu Ala 145 150 155 160 Ala Tyr Gly Thr Ile Val Ser Ala Glu Leu Thr Gly Arg His Lys Gly 165 170 175 Pro Leu Ala Glu Arg Asp Phe Lys Ser Gly Arg Cys Pro Gly Asn Ser 180 185 190 Phe Ser Cys Gly Asn Ser Gln Cys Val Thr Lys Val Asn Pro Glu Cys 195 200 205 Asp Asp Gln Glu Asp Cys Ser Asp Gly Ser Asp Glu Ala His Cys Glu 210 215 220 Cys Gly Leu Gln Pro Ala Trp Arg Met Ala Gly Arg Ile Val Gly Gly 225 230 235 240 Met Glu Ala Ser Pro Gly Glu Phe Pro Trp Gln Ala Ser Leu Arg Glu 245 250 255 Asn Lys Glu His Phe Cys Gly Ala Ala Ile Ile Asn Ala Arg Trp Leu 260 265 270 Val Ser Ala Ala His Cys Phe Asn Glu Phe Gln Asp Pro Thr Lys Trp 275 280 285 Val Ala Tyr Val Gly Ala Thr Tyr Leu Ser Gly Ser Glu Ala Ser Thr 290 295 300 Val Arg Ala Gln Val Val Gln Ile Val Lys His Pro Leu Tyr Asn Ala 305 310 315 320 Asp Thr Ala Asp Phe Asp Val Ala Val Leu Glu Leu Thr Ser Pro Leu 325 330 335 Pro Phe Gly Arg His Ile Gln Pro Val Cys Leu Pro Ala Ala Thr His 340 345 350 Ile Phe Pro Pro Ser Lys Lys Cys Leu Ile Ser Gly Trp Gly Tyr Leu 355 360 365 Lys Glu Asp Phe Leu Val Lys Pro Gly Val Leu Gln Lys Ala Thr Val 370 375 380 Glu Leu Leu Asp Gln Ala Leu Cys Ala Ser Leu Tyr Gly His Ser Leu 385 390 395 400 Thr Asp Arg Met Val Cys Ala Gly Tyr Leu Asp Gly Lys Val Asp Ser 405 410 415 Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Cys Glu Glu Pro Ser Gly 420 425 430 Arg Phe Ser Leu Ala Gly Ile Val Ser Trp Gly Ile Gly Cys Ala Glu 435 440 445 Ala Arg Arg Pro Gly Val Tyr Ala Arg Val Thr Arg Leu Arg Asp Trp 450 455 460 Ile Leu Glu Ala Thr Thr Lys Ala Ser Met Pro Leu Ala Pro Thr Met 465 470 475 480 Ala Pro Ala Pro Ala Ala Pro Ser Thr Ala Trp Pro Thr Ser Pro Glu 485 490 495 Ser Pro Val Val Ser Thr Pro Thr Lys Ser Met Gln Ala Leu Ser Thr 500 505 510 Val Pro Leu Asp Trp Val Thr Val Pro Lys Leu Gln Glu Cys Gly Ala 515 520 525 Arg Pro Ala Met Glu Lys Pro Thr Arg Val Val Gly Gly Phe Gly Ala 530 535 540 Ala Ser Gly Glu Val Pro Trp Gln Val Ser Leu Lys Glu Gly Ser Arg 545 550 555 560 His Phe Cys Gly Ala Thr Val Val Gly Asp Arg Trp Leu Leu Ser Ala 565 570 575 Ala His Cys Phe Asn His Thr Lys Val Glu Gln Val Arg Ala His Leu 580 585 590 Gly Thr Ala Ser Leu Leu Gly Leu Gly Gly Ser Pro Val Lys Ile Gly 595 600 605 Leu Arg Arg Val Val Leu His Pro Leu Tyr Asn Pro Gly Ile Leu Asp 610 615 620 Phe Asp Leu Ala Val Leu Glu Leu Ala Ser Pro Leu Ala Phe Asn Lys 625 630 635 640 Tyr Ile Gln Pro Val Cys Leu Pro Leu Ala Ile Arg Lys Phe Pro Val 645 650 655 Gly Arg Lys Cys Met Ile Ser Gly Trp Gly Asn Thr Gln Glu Gly Asn 660 665 670 Ala Thr Lys Pro Glu Leu Leu Gln Lys Ala Ser Val Gly Ile Ile Asp 675 680 685 Gln Lys Thr Cys Ser Val Leu Tyr Asn Phe Ser Leu Thr Asp Arg Met 690 695 700 Ile Cys Ala Gly Phe Leu Glu Gly Lys Val Asp Ser Cys Gln Gly Asp 705 710 715 720 Ser Gly Gly Pro Leu Ala Cys Glu Glu Ala Pro Gly Val Phe Tyr Leu 725 730 735 Ala Gly Ile Val Ser Trp Gly Ile Gly Cys Ala Gln Val Lys Lys Pro 740 745 750 Gly Val Tyr Thr Arg Ile Thr Arg Leu Lys Gly Trp Ile Leu Glu Ile 755 760 765 Met Ser Ser Gln Pro Leu Pro Met Ser Pro Pro Ser Thr Thr Arg Met 770 775 780 Leu Ala Thr Thr Ser Pro Arg Thr Thr Ala Gly Leu Thr Val Pro Gly 785 790 795 800 Ala Thr Pro Ser Arg Pro Thr Pro Gly Ala Ala Ser Arg Val Thr Gly 805 810 815 Gln Pro Ala Asn Ser Thr Leu Ser Ala Val Ser Thr Thr Ala Arg Gly 820 825 830 Gln Thr Pro Phe Pro Asp Ala Pro Glu Ala Thr Thr His Thr Gln Leu 835 840 845 Pro Asp Cys Gly Leu Ala Pro Ala Ala Leu Thr Arg Ile Val Gly Gly 850 855 860 Ser Ala Ala Gly Arg Gly Glu Trp Pro Trp Gln Val Gly Leu Trp Leu 865 870 875 880 Arg Arg Arg Glu His Arg Cys Gly Ala Val Leu Val Ala Glu Arg Trp 885 890 895 Leu Leu Ser Ala Ala His Cys Phe Asp Val Tyr Gly Asp Pro Lys Gln 900 905 910 Trp Ala Ala Phe Leu Gly Thr Pro Phe Leu Ser Gly Ala Glu Gly Gln 915 920 925 Leu Glu Arg Val Ala Arg Ile Tyr Lys His Pro Phe Tyr Asn Leu Tyr 930 935 940 Thr Leu Asp Tyr Asp Val Ala Leu Leu Glu Leu Ala Gly Pro Val Arg 945 950 955 960 Arg Ser Arg Leu Val Arg Pro Ile Cys Leu Pro Glu Pro Ala Pro Arg 965 970 975 Pro Pro Asp Gly Thr Arg Cys Val Ile Thr Gly Trp Gly Ser Val Arg 980 985 990 Glu Gly Gly Ser Met Ala Arg Gln Leu Gln Lys Ala Ala Val Arg Leu 995 1000 1005 Leu Ser Glu Gln Thr Cys Arg Arg Phe Tyr Pro Val Gln Ile Ser Ser 1010 1015 1020 Arg Met Leu Cys Ala Gly Phe Pro Gln Gly Gly Val Asp Ser Cys Ser 1025 1030 1035 1040 Gly Asp Ala Gly Gly Pro Leu Ala Cys Arg Glu Pro Ser Gly Arg Trp 1045 1050 1055 Val Leu Thr Gly Val Thr Ser Trp Gly Tyr Gly Cys Gly Arg Pro His 1060 1065 1070 Phe Pro Gly Val Tyr Thr Arg Val Ala Ala Val Arg Gly Trp Ile Gly 1075 1080 1085 Gln His Ile Gln Glu 1090 <210> SEQ ID NO 21 <211> LENGTH: 702 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (1)...(699) <223> OTHER INFORMATION: Nucleic Acid encoding protease domain of endotheliase 1 <400> SEQUENCE: 21 agg atc gtt ggt ggg aca gaa gta gaa gag ggt gaa tgg ccc tgg cag 48 Arg Ile Val Gly Gly Thr Glu Val Glu Glu Gly Glu Trp Pro Trp Gln 1 5 10 15 gct agc ctg cag tgg gat ggg agt cat cgc tgt gga gca acc tta att 96 Ala Ser Leu Gln Trp Asp Gly Ser His Arg Cys Gly Ala Thr Leu Ile 20 25 30 aat gcc aca tgg ctt gtg agt gct gct cac tgt ttt aca aca tat aag 144 Asn Ala Thr Trp Leu Val Ser Ala Ala His Cys Phe Thr Thr Tyr Lys 35 40 45 aac cct gcc aga tgg act gct tcc ttt gga gta aca ata aaa cct tcg 192 Asn Pro Ala Arg Trp Thr Ala Ser Phe Gly Val Thr Ile Lys Pro Ser 50 55 60 aaa atg aaa cgg ggt ctc cgg aga ata att gtc cat gaa aaa tac aaa 240 Lys Met Lys Arg Gly Leu Arg Arg Ile Ile Val His Glu Lys Tyr Lys 65 70 75 80 cac cca tca cat gac tat gat att tct ctt gca gag ctt tct agc cct 288His Pro Ser His Asp Tyr Asp Ile Ser Leu Ala Glu Leu Ser Ser Pro 85 90 95 gtt ccc tac aca aat gca gta cat aga gtt tgt ctc cct gat gca tcc 336Val Pro Tyr Thr Asn Ala Val His Arg Val Cys Leu Pro Asp Ala Ser 100 105 110 tat gag ttt caa cca ggt gat gtg atg ttt gtg aca gga ttt gga gca 384Tyr Glu Phe Gln Pro Gly Asp Val Met Phe Val Thr Gly Phe Gly Ala 115 120 125 ctg aaa aat gat ggt tac agt caa aat cat ctt cga caa gca cag gtg 432Leu Lys Asn Asp Gly Tyr Ser Gln Asn His Leu Arg Gln Ala Gln Val 130 135 140 act ctc ata gac gct aca act tgc aat gaa cct caa gct tac aat gac 480 Thr Leu Ile Asp Ala Thr Thr Cys Asn Glu Pro Gln Ala Tyr Asn Asp 145 150 155 160 gcc ata act cct aga atg tta tgt gct ggc tcc tta gaa gga aaa aca 528 Ala Ile Thr Pro Arg Met Leu Cys Ala Gly Ser Leu Glu Gly Lys Thr 165 170 175 gat gca tgc cag ggt gac tct gga gga cca ctg gtt agt tca gat gct 576 Asp Ala Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Ser Ser Asp Ala 180 185 190 aga gat atc tgg tac ctt gct gga ata gtg agc tgg gga gat gaa tgt 624 Arg Asp Ile Trp Tyr Leu Ala Gly Ile Val Ser Trp Gly Asp Glu Cys 195 200 205 gcg aaa ccc aac aag cct ggt gtt tat act aga gtt acg gcc ttg cgg 672 Ala Lys Pro Asn Lys Pro Gly Val Tyr Thr Arg Val Thr Ala Leu Arg 210 215 220 gac tgg att act tca aaa act ggt atc taa 702 Asp Trp Ile Thr Ser Lys Thr Gly Ile 225 230 <210> SEQ ID NO 22 <211> LENGTH: 233 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: SITE <222> LOCATION: (1)...(233) <223> OTHER INFORMATION: Protease domain of endotheliase 1 <400> SEQUENCE: 22 Arg Ile Val Gly Gly Thr Glu Val Glu Glu Gly Glu Trp Pro Trp Gln 1 5 10 15 Ala Ser Leu Gln Trp Asp Gly Ser His Arg Cys Gly Ala Thr Leu Ile 20 25 30 Asn Ala Thr Trp Leu Val Ser Ala Ala His Cys Phe Thr Thr Tyr Lys 35 40 45 Asn Pro Ala Arg Trp Thr Ala Ser Phe Gly Val Thr Ile Lys Pro Ser 50 55 60 Lys Met Lys Arg Gly Leu Arg Arg Ile Ile Val His Glu Lys Tyr Lys 65 70 75 80 His Pro Ser His Asp Tyr Asp Ile Ser Leu Ala Glu Leu Ser Ser Pro 85 90 95 Val Pro Tyr Thr Asn Ala Val His Arg Val Cys Leu Pro Asp Ala Ser 100 105 110 Tyr Glu Phe Gln Pro Gly Asp Val Met Phe Val Thr Gly Phe Gly Ala 115 120 125 Leu Lys Asn Asp Gly Tyr Ser Gln Asn His Leu Arg Gln Ala Gln Val 130 135 140 Thr Leu Ile Asp Ala Thr Thr Cys Asn Glu Pro Gln Ala Tyr Asn Asp 145 150 155 160 Ala Ile Thr Pro Arg Met Leu Cys Ala Gly Ser Leu Glu Gly Lys Thr 165 170 175 Asp Ala Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Ser Ser Asp Ala 180 185 190 Arg Asp Ile Trp Tyr Leu Ala Gly Ile Val Ser Trp Gly Asp Glu Cys 195 200 205 Ala Lys Pro Asn Lys Pro Gly Val Tyr Thr Arg Val Thr Ala Leu Arg 210 215 220 Asp Trp Ile Thr Ser Lys Thr Gly Ile 225 230 <210> SEQ ID NO 23 <211> LENGTH: 1689 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (1)...(1689) <223> OTHER INFORMATION: Nucleic acid encoding Endotheliase 2-S protein <400> SEQUENCE: 23 atg gag agg gac agc cac ggg aat gca tct cca gca aga aca cct tca 48 Met Glu Arg Asp Ser His Gly Asn Ala Ser Pro Ala Arg Thr Pro Ser 1 5 10 15 gct gga gca tct cca gcc cag gca tct cca gct ggg aca cct cca ggc 96 Ala Gly Ala Ser Pro Ala Gln Ala Ser Pro Ala Gly Thr Pro Pro Gly 20 25 30 cgg gca tct cca gcc cag gca tct cca gcc cag gca tct cca gct ggg 144 Arg Ala Ser Pro Ala Gln Ala Ser Pro Ala Gln Ala Ser Pro Ala Gly 35 40 45 aca cct ccg ggc cgg gca tct cca gcc cag gca tct cca gct ggt aca 192 Thr Pro Pro Gly Arg Ala Ser Pro Ala Gln Ala Ser Pro Ala Gly Thr 50 55 60 cct cca ggc cgg gca tct cca ggc cgg gca tct cca gcc cag gca tct 240 Pro Pro Gly Arg Ala Ser Pro Gly Arg Ala Ser Pro Ala Gln Ala Ser 65 70 75 80 cca gcc cgg gca tct ccg gct ctg gca tca ctt tcc agg tcc tca tcc 288Pro Ala Arg Ala Ser Pro Ala Leu Ala Ser Leu Ser Arg Ser Ser Ser 85 90 95 ggc agg tca tca tcc gcc agg tca gcc tcg gtg aca acc tcc cca acc 336Gly Arg Ser Ser Ser Ala Arg Ser Ala Ser Val Thr Thr Ser Pro Thr 100 105 110 aga gtg tac ctt gtt aga gca aca cca gtg ggg gct gta ccc atc cga 384Arg Val Tyr Leu Val Arg Ala Thr Pro Val Gly Ala Val Pro Ile Arg 115 120 125 tca tct cct gcc agg tca gca cca gca acc agg gcc acc agg gag agc 432 Ser Ser Pro Ala Arg Ser Ala Pro Ala Thr Arg Ala Thr Arg Glu Ser 130 135 140 cca ggt acg agc ctg ccc aag ttc acc tgg cgg gag ggc cag aag cag 480Pro Gly Thr Ser Leu Pro Lys Phe Thr Trp Arg Glu Gly Gln Lys Gln 145 150 155 160 cta ccg ctc atc ggg tgc gtg ctc ctc ctc att gcc ctg gtg gtt tcg 528Leu Pro Leu Ile Gly Cys Val Leu Leu Leu Ile Ala Leu Val Val Ser 165 170 175 ctc atc atc ctc ttc cag ttc tgg cag ggc cac aca ggg atc agg tac 576 Leu Ile Ile Leu Phe Gln Phe Trp Gln Gly His Thr Gly Ile Arg Tyr 180 185 190 aag gag cag agg gag agc tgt ccc aag cac gct gtt cgc tgt gac ggg 624 Lys Glu Gln Arg Glu Ser Cys Pro Lys His Ala Val Arg Cys Asp Gly 195 200 205 gtg gtg gac tgc aag ctg aag agt gac gag ctg ggc tgc gtg agg ttt 672 Val Val Asp Cys Lys Leu Lys Ser Asp Glu Leu Gly Cys Val Arg Phe 210 215 220 gac tgg gac aag tct ctg ctt aaa atc tac tct ggg tcc tcc cat cag 720 Asp Trp Asp Lys Ser Leu Leu Lys Ile Tyr Ser Gly Ser Ser His Gln 225 230 235 240 tgg ctt ccc atc tgt agc agc aac tgg aat gac tcc tac tca gag aag 768 Trp Leu Pro Ile Cys Ser Ser Asn Trp Asn Asp Ser Tyr Ser Glu Lys 245 250 255 acc tgc cag cag ctg ggt ttc gag agt gct cac cgg aca acc gag gtt 816 Thr Cys Gln Gln Leu Gly Phe Glu Ser Ala His Arg Thr Thr Glu Val 260 265 270 gcc cac agg gat ttt gcc aac agc ttc tca atc ttg aga tac aac tcc 864 Ala His Arg Asp Phe Ala Asn Ser Phe Ser Ile Leu Arg Tyr Asn Ser 275 280 285 acc atc cag gaa agc ctc cac agg tct gaa tgc cct tcc cag cgg tat 912 Thr Ile Gln Glu Ser Leu His Arg Ser Glu Cys Pro Ser Gln Arg Tyr 290 295 300 atc tcc ctc cag tgt tcc cac tgc gga ctg agg gcc atg acc ggg cgg 960 Ile Ser Leu Gln Cys Ser His Cys Gly Leu Arg Ala Met Thr Gly Arg 305 310 315 320 atc gtg gga ggg gcg ctg gcc tcg gat agc aag tgg cct tgg caa gtg 1008 Ile Val Gly Gly Ala Leu Ala Ser Asp Ser Lys Trp Pro Trp Gln Val 325 330 335 agt ctg cac ttc ggc acc acc cac atc tgt gga ggc acg ctc att gac 1056 Ser Leu His Phe Gly Thr Thr His Ile Cys Gly Gly Thr Leu Ile Asp 340 345 350 gcc cag tgg gtg ctc act gcc gcc cac tgc ttc ttc gtg acc cgg gag 1104 Ala Gln Trp Val Leu Thr Ala Ala His Cys Phe Phe Val Thr Arg Glu 355 360 365 aag gtc ctg gag ggc tgg aag gtg tac gcg ggc acc agc aac ctg cac 1152 Lys Val Leu Glu Gly Trp Lys Val Tyr Ala Gly Thr Ser Asn Leu His 370 375 380 cag ttg cct gag gca gcc tcc att gcc gag atc atc atc aac agc aat 1200 Gln Leu Pro Glu Ala Ala Ser Ile Ala Glu Ile Ile Ile Asn Ser Asn 385 390 395 400 tac acc gat gag gag gac gac tat gac atc gcc ctc atg cgg ctg tcc 1248 Tyr Thr Asp Glu Glu Asp Asp Tyr Asp Ile Ala Leu Met Arg Leu Ser 405 410 415 aag ccc ctg acc ctg tcc gct cac atc cac cct gct tgc ctc ccc atg 1296 Lys Pro Leu Thr Leu Ser Ala His Ile His Pro Ala Cys Leu Pro Met 420 425 430 cat gga cag acc ttt agc ctc aat gag acc tgc tgg atc aca ggc ttt 1344 His Gly Gln Thr Phe Ser Leu Asn Glu Thr Cys Trp Ile Thr Gly Phe 435 440 445 ggc aag acc agg gag aca gat gac aag aca tcc ccc ttc ctc cgg gag 1392 Gly Lys Thr Arg Glu Thr Asp Asp Lys Thr Ser Pro Phe Leu Arg Glu 450 455 460 gtg cag gtc aat ctc atc gac ttc aag aaa tgc aat gac tac ttg gtc 1440 Val Gln Val Asn Leu Ile Asp Phe Lys Lys Cys Asn Asp Tyr Leu Val 465 470 475 480 tat gac agt tac ctt acc cca agg atg atg tgt gct ggg gac ctt cgt 1488 Tyr Asp Ser Tyr Leu Thr Pro Arg Met Met Cys Ala Gly Asp Leu Arg 485 490 495 ggg ggc aga gac tcc tgc cag gga gac agc ggg ggg cct ctt gtc tgt 1536 Gly Gly Arg Asp Ser Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Cys 500 505 510 gag cag aac aac cgc tgg tac ctg gca ggt gtc acc agc tgg ggc aca 1584 Glu Gln Asn Asn Arg Trp Tyr Leu Ala Gly Val Thr Ser Trp Gly Thr 515 520 525 ggc tgt ggc cag aga aac aaa cct ggt gtg tac acc aaa gtg aca gaa 1632 Gly Cys Gly Gln Arg Asn Lys Pro Gly Val Tyr Thr Lys Val Thr Glu 530 535 540 gtt ctt ccc tgg att tac agc aag atg gag agc gag gtg cga ttc ata 1680 Val Leu Pro Trp Ile Tyr Ser Lys Met Glu Ser Glu Val Arg Phe Ile 545 550 555 560 aaa tcc taa 1689 Lys Ser * <210> SEQ ID NO 24 <211> LENGTH: 562 <212> TYPE: PRT <213> ORGANISM: homo sapien <220> FEATURE: <221> NAME/KEY: protease domain of endotheliase 2 <222> LOCATION: (321)..(562) <400> SEQUENCE: 24 Met Glu Arg Asp Ser His Gly Asn Ala Ser Pro Ala Arg Thr Pro Ser 1 5 10 15 Ala Gly Ala Ser Pro Ala Gln Ala Ser Pro Ala Gly Thr Pro Pro Gly 20 25 30 Arg Ala Ser Pro Ala Gln Ala Ser Pro Ala Gln Ala Ser Pro Ala Gly 35 40 45 Thr Pro Pro Gly Arg Ala Ser Pro Ala Gln Ala Ser Pro Ala Gly Thr 50 55 60 Pro Pro Gly Arg Ala Ser Pro Gly Arg Ala Ser Pro Ala Gln Ala Ser 65 70 75 80 Pro Ala Arg Ala Ser Pro Ala Leu Ala Ser Leu Ser Arg Ser Ser Ser 85 90 95 Gly Arg Ser Ser Ser Ala Arg Ser Ala Ser Val Thr Thr Ser Pro Thr 100 105 110 Arg Val Tyr Leu Val Arg Ala Thr Pro Val Gly Ala Val Pro Ile Arg 115 120 125 Ser Ser Pro Ala Arg Ser Ala Pro Ala Thr Arg Ala Thr Arg Glu Ser 130 135 140 Pro Gly Thr Ser Leu Pro Lys Phe Thr Trp Arg Glu Gly Gln Lys Gln 145 150 155 160 Leu Pro Leu Ile Gly Cys Val Leu Leu Leu Ile Ala Leu Val Val Ser 165 170 175 Leu Ile Ile Leu Phe Gln Phe Trp Gln Gly His Thr Gly Ile Arg Tyr 180 185 190 Lys Glu Gln Arg Glu Ser Cys Pro Lys His Ala Val Arg Cys Asp Gly 195 200 205 Val Val Asp Cys Lys Leu Lys Ser Asp Glu Leu Gly Cys Val Arg Phe 210 215 220 Asp Trp Asp Lys Ser Leu Leu Lys Ile Tyr Ser Gly Ser Ser His Gln 225 230 235 240 Trp Leu Pro Ile Cys Ser Ser Asn Trp Asn Asp Ser Tyr Ser Glu Lys 245 250 255 Thr Cys Gln Gln Leu Gly Phe Glu Ser Ala His Arg Thr Thr Glu Val 260 265 270 Ala His Arg Asp Phe Ala Asn Ser Phe Ser Ile Leu Arg Tyr Asn Ser 275 280 285 Thr Ile Gln Glu Ser Leu His Arg Ser Glu Cys Pro Ser Gln Arg Tyr 290 295 300 Ile Ser Leu Gln Cys Ser His Cys Gly Leu Arg Ala Met Thr Gly Arg 305 310 315 320 Ile Val Gly Gly Ala Leu Ala Ser Asp Ser Lys Trp Pro Trp Gln Val 325 330 335 Ser Leu His Phe Gly Thr Thr His Ile Cys Gly Gly Thr Leu Ile Asp 340 345 350 Ala Gln Trp Val Leu Thr Ala Ala His Cys Phe Phe Val Thr Arg Glu 355 360 365 Lys Val Leu Glu Gly Trp Lys Val Tyr Ala Gly Thr Ser Asn Leu His 370 375 380 Gln Leu Pro Glu Ala Ala Ser Ile Ala Glu Ile Ile Ile Asn Ser Asn 385 390 395 400 Tyr Thr Asp Glu Glu Asp Asp Tyr Asp Ile Ala Leu Met Arg Leu Ser 405 410 415 Lys Pro Leu Thr Leu Ser Ala His Ile His Pro Ala Cys Leu Pro Met 420 425 430 His Gly Gln Thr Phe Ser Leu Asn Glu Thr Cys Trp Ile Thr Gly Phe 435 440 445 Gly Lys Thr Arg Glu Thr Asp Asp Lys Thr Ser Pro Phe Leu Arg Glu 450 455 460 Val Gln Val Asn Leu Ile Asp Phe Lys Lys Cys Asn Asp Tyr Leu Val 465 470 475 480 Tyr Asp Ser Tyr Leu Thr Pro Arg Met Met Cys Ala Gly Asp Leu Arg 485 490 495 Gly Gly Arg Asp Ser Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Cys 500 505 510 Glu Gln Asn Asn Arg Trp Tyr Leu Ala Gly Val Thr Ser Trp Gly Thr 515 520 525 Gly Cys Gly Gln Arg Asn Lys Pro Gly Val Tyr Thr Lys Val Thr Glu 530 535 540 Val Leu Pro Trp Ile Tyr Ser Lys Met Glu Ser Glu Val Arg Phe Ile 545 550 555 560 Lys Ser <210> SEQ ID NO 25 <211> LENGTH: 2067 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (1)...(2067) <223> OTHER INFORMATION: Nucleic acid encoding (endotheliase 2-L) protein <400> SEQUENCE: 25 atg gag agg gac agc cac ggg aat gca tct cca gca aga aca cct tca 48 Met Glu Arg Asp Ser His Gly Asn Ala Ser Pro Ala Arg Thr Pro Ser 1 5 10 15 gct gga gca tct cca gcc cag gca tct cca gct ggg aca cct cca ggc 96 Ala Gly Ala Ser Pro Ala Gln Ala Ser Pro Ala Gly Thr Pro Pro Gly 20 25 30 cgg gca tct cca gcc cag gca tct cca gcc cag gca tct cca gct ggg 144 Arg Ala Ser Pro Ala Gln Ala Ser Pro Ala Gln Ala Ser Pro Ala Gly 35 40 45 aca cct ccg ggc cgg gca tct cca gcc cag gca tct cca gct ggt aca 192 Thr Pro Pro Gly Arg Ala Ser Pro Ala Gln Ala Ser Pro Ala Gly Thr 50 55 60 cct cca ggc cgg gca tct cca ggc cgg gca tct cca gcc cag gca tct 240 Pro Pro Gly Arg Ala Ser Pro Gly Arg Ala Ser Pro Ala Gln Ala Ser 65 70 75 80 cca gcc cgg gca tct ccg gct ctg gca tca ctt tcc agg tcc tca tcc 288Pro Ala Arg Ala Ser Pro Ala Leu Ala Ser Leu Ser Arg Ser Ser Ser 85 90 95 ggc agg tca tca tcc gcc agg tca gcc tcg gtg aca acc tcc cca acc 336Gly Arg Ser Ser Ser Ala Arg Ser Ala Ser Val Thr Thr Ser Pro Thr 100 105 110 aga gtg tac ctt gtt aga gca aca cca gtg ggg gct gta ccc atc cga 384Arg Val Tyr Leu Val Arg Ala Thr Pro Val Gly Ala Val Pro Ile Arg 115 120 125 tca tct cct gcc agg tca gca cca gca acc agg gcc acc agg gag agc 432 Ser Ser Pro Ala Arg Ser Ala Pro Ala Thr Arg Ala Thr Arg Glu Ser 130 135 140 cca ggt acg agc ctg ccc aag ttc acc tgg cgg gag ggc cag aag cag 480Pro Gly Thr Ser Leu Pro Lys Phe Thr Trp Arg Glu Gly Gln Lys Gln 145 150 155 160 cta ccg ctc atc ggg tgc gtg ctc ctc ctc att gcc ctg gtg gtt tcg 528Leu Pro Leu Ile Gly Cys Val Leu Leu Leu Ile Ala Leu Val Val Ser 165 170 175 ctc atc atc ctc ttc cag ttc tgg cag ggc cac aca ggg atc agg tac 576 Leu Ile Ile Leu Phe Gln Phe Trp Gln Gly His Thr Gly Ile Arg Tyr 180 185 190 aag gag cag agg gag agc tgt ccc aag cac gct gtt cgc tgt gac ggg 624 Lys Glu Gln Arg Glu Ser Cys Pro Lys His Ala Val Arg Cys Asp Gly 195 200 205 gtg gtg gac tgc aag ctg aag agt gac gag ctg ggc tgc gtg agg ttt 672 Val Val Asp Cys Lys Leu Lys Ser Asp Glu Leu Gly Cys Val Arg Phe 210 215 220 gac tgg gac aag tct ctg ctt aaa atc tac tct ggg tcc tcc cat cag 720 Asp Trp Asp Lys Ser Leu Leu Lys Ile Tyr Ser Gly Ser Ser His Gln 225 230 235 240 tgg ctt ccc atc tgt agc agc aac tgg aat gac tcc tac tca gag aag 768 Trp Leu Pro Ile Cys Ser Ser Asn Trp Asn Asp Ser Tyr Ser Glu Lys 245 250 255 acc tgc cag cag ctg ggt ttc gag agt gct cac cgg aca acc gag gtt 816 Thr Cys Gln Gln Leu Gly Phe Glu Ser Ala His Arg Thr Thr Glu Val 260 265 270 gcc cac agg gat ttt gcc aac agc ttc tca atc ttg aga tac aac tcc 864 Ala His Arg Asp Phe Ala Asn Ser Phe Ser Ile Leu Arg Tyr Asn Ser 275 280 285 acc atc cag gaa agc ctc cac agg tct gaa tgc cct tcc cag cgg tat 912 Thr Ile Gln Glu Ser Leu His Arg Ser Glu Cys Pro Ser Gln Arg Tyr 290 295 300 atc tcc ctc cag tgt tcc cac tgc gga ctg agg gcc atg acc ggg cgg 960 Ile Ser Leu Gln Cys Ser His Cys Gly Leu Arg Ala Met Thr Gly Arg 305 310 315 320 atc gtg gga ggg gcg ctg gcc tcg gat agc aag tgg cct tgg caa gtg 1008 Ile Val Gly Gly Ala Leu Ala Ser Asp Ser Lys Trp Pro Trp Gln Val 325 330 335 agt ctg cac ttc ggc acc acc cac atc tgt gga ggc acg ctc att gac 1056 Ser Leu His Phe Gly Thr Thr His Ile Cys Gly Gly Thr Leu Ile Asp 340 345 350 gcc cag tgg gtg ctc act gcc gcc cac tgc ttc ttc gtg acc cgg gag 1104 Ala Gln Trp Val Leu Thr Ala Ala His Cys Phe Phe Val Thr Arg Glu 355 360 365 aag gtc ctg gag ggc tgg aag gtg tac gcg ggc acc agc aac ctg cac 1152 Lys Val Leu Glu Gly Trp Lys Val Tyr Ala Gly Thr Ser Asn Leu His 370 375 380 cag ttg cct gag gca gcc tcc att gcc gag atc atc atc aac agc aat 1200 Gln Leu Pro Glu Ala Ala Ser Ile Ala Glu Ile Ile Ile Asn Ser Asn 385 390 395 400 tac acc gat gag gag gac gac tat gac atc gcc ctc atg cgg ctg tcc 1248 Tyr Thr Asp Glu Glu Asp Asp Tyr Asp Ile Ala Leu Met Arg Leu Ser 405 410 415 aag ccc ctg acc ctg tcc gct cac atc cac cct gct tgc ctc ccc atg 1296 Lys Pro Leu Thr Leu Ser Ala His Ile His Pro Ala Cys Leu Pro Met 420 425 430 cat gga cag acc ttt agc ctc aat gag acc tgc tgg atc aca ggc ttt 1344 His Gly Gln Thr Phe Ser Leu Asn Glu Thr Cys Trp Ile Thr Gly Phe 435 440 445 ggc aag acc agg gag aca gat gac aag aca tcc ccc ttc ctc cgg gag 1392 Gly Lys Thr Arg Glu Thr Asp Asp Lys Thr Ser Pro Phe Leu Arg Glu 450 455 460 gtg cag gtc aat ctc atc gac ttc aag aaa tgc aat gac tac ttg gtc 1440 Val Gln Val Asn Leu Ile Asp Phe Lys Lys Cys Asn Asp Tyr Leu Val 465 470 475 480 tat gac agt tac ctt acc cca agg atg atg tgt gct ggg gac ctt cgt 1488 Tyr Asp Ser Tyr Leu Thr Pro Arg Met Met Cys Ala Gly Asp Leu Arg 485 490 495 ggg ggc aga gac tcc tgc cag gga gac agc ggg ggg cct ctt gtc tgt 1536 Gly Gly Arg Asp Ser Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Cys 500 505 510 gag cag aac aac cgc tgg tac ctg gca ggt gtc acc agc tgg ggc aca 1584 Glu Gln Asn Asn Arg Trp Tyr Leu Ala Gly Val Thr Ser Trp Gly Thr 515 520 525 ggc tgt ggc cag aga aac aaa cct ggt gtg tac acc aaa gtg aca gaa 1632 Gly Cys Gly Gln Arg Asn Lys Pro Gly Val Tyr Thr Lys Val Thr Glu 530 535 540 gtt ctt ccc tgg att tac agc aag atg gag aac aga gct cag cgg gtt 1680 Val Leu Pro Trp Ile Tyr Ser Lys Met Glu Asn Arg Ala Gln Arg Val 545 550 555 560 gaa aaa gcg tgg acc tac agg cca ggc agg cag ttg ctg ggc aga tgt 1728 Glu Lys Ala Trp Thr Tyr Arg Pro Gly Arg Gln Leu Leu Gly Arg Cys 565 570 575 tct ccc aga agt att ttt ttg tgt aag gtt gca atg gac ttt gaa aac 1776 Ser Pro Arg Ser Ile Phe Leu Cys Lys Val Ala Met Asp Phe Glu Asn 580 585 590 gtt tca gtt tct gca gag gat ttt gtg ata gtt ttt gtt atc aag cat 1824 Val Ser Val Ser Ala Glu Asp Phe Val Ile Val Phe Val Ile Lys His 595 600 605 tta tgc atg gga atc cgc tct tca tgg cct ttc cca gct ctg ttt gtt 1872 Leu Cys Met Gly Ile Arg Ser Ser Trp Pro Phe Pro Ala Leu Phe Val 610 615 620 tta gtc ttt ttg att ttc ttt ttg ttg ttg ttg ttg tct ttt tta aaa 1920 Leu Val Phe Leu Ile Phe Phe Leu Leu Leu Leu Leu Ser Phe Leu Lys 625 630 635 640 aac aca agt gac tcc att ttg act ctg aca act ttc aca gct gtc acc 1968 Asn Thr Ser Asp Ser Ile Leu Thr Leu Thr Thr Phe Thr Ala Val Thr 645 650 655 aga atg ctc cct gag aac tac cat tct ttc cct ttc cca ctt aaa ata 2016 Arg Met Leu Pro Glu Asn Tyr His Ser Phe Pro Phe Pro Leu Lys Ile 660 665 670 ttt cat cag aac ctc act act atc ata aaa gag tat aaa gta ata aaa 2064 Phe His Gln Asn Leu Thr Thr Ile Ile Lys Glu Tyr Lys Val Ile Lys 675 680 685 taa 2067 <210> SEQ ID NO 26 <211> LENGTH: 688 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: protease domain <222> LOCATION: (321)..(688) <400> SEQUENCE: 26 Met Glu Arg Asp Ser His Gly Asn Ala Ser Pro Ala Arg Thr Pro Ser 1 5 10 15 Ala Gly Ala Ser Pro Ala Gln Ala Ser Pro Ala Gly Thr Pro Pro Gly 20 25 30 Arg Ala Ser Pro Ala Gln Ala Ser Pro Ala Gln Ala Ser Pro Ala Gly 35 40 45 Thr Pro Pro Gly Arg Ala Ser Pro Ala Gln Ala Ser Pro Ala Gly Thr 50 55 60 Pro Pro Gly Arg Ala Ser Pro Gly Arg Ala Ser Pro Ala Gln Ala Ser 65 70 75 80 Pro Ala Arg Ala Ser Pro Ala Leu Ala Ser Leu Ser Arg Ser Ser Ser 85 90 95 Gly Arg Ser Ser Ser Ala Arg Ser Ala Ser Val Thr Thr Ser Pro Thr 100 105 110 Arg Val Tyr Leu Val Arg Ala Thr Pro Val Gly Ala Val Pro Ile Arg 115 120 125 Ser Ser Pro Ala Arg Ser Ala Pro Ala Thr Arg Ala Thr Arg Glu Ser 130 135 140 Pro Gly Thr Ser Leu Pro Lys Phe Thr Trp Arg Glu Gly Gln Lys Gln 145 150 155 160 Leu Pro Leu Ile Gly Cys Val Leu Leu Leu Ile Ala Leu Val Val Ser 165 170 175 Leu Ile Ile Leu Phe Gln Phe Trp Gln Gly His Thr Gly Ile Arg Tyr 180 185 190 Lys Glu Gln Arg Glu Ser Cys Pro Lys His Ala Val Arg Cys Asp Gly 195 200 205 Val Val Asp Cys Lys Leu Lys Ser Asp Glu Leu Gly Cys Val Arg Phe 210 215 220 Asp Trp Asp Lys Ser Leu Leu Lys Ile Tyr Ser Gly Ser Ser His Gln 225 230 235 240 Trp Leu Pro Ile Cys Ser Ser Asn Trp Asn Asp Ser Tyr Ser Glu Lys 245 250 255 Thr Cys Gln Gln Leu Gly Phe Glu Ser Ala His Arg Thr Thr Glu Val 260 265 270 Ala His Arg Asp Phe Ala Asn Ser Phe Ser Ile Leu Arg Tyr Asn Ser 275 280 285 Thr Ile Gln Glu Ser Leu His Arg Ser Glu Cys Pro Ser Gln Arg Tyr 290 295 300 Ile Ser Leu Gln Cys Ser His Cys Gly Leu Arg Ala Met Thr Gly Arg 305 310 315 320 Ile Val Gly Gly Ala Leu Ala Ser Asp Ser Lys Trp Pro Trp Gln Val 325 330 335 Ser Leu His Phe Gly Thr Thr His Ile Cys Gly Gly Thr Leu Ile Asp 340 345 350 Ala Gln Trp Val Leu Thr Ala Ala His Cys Phe Phe Val Thr Arg Glu 355 360 365 Lys Val Leu Glu Gly Trp Lys Val Tyr Ala Gly Thr Ser Asn Leu His 370 375 380 Gln Leu Pro Glu Ala Ala Ser Ile Ala Glu Ile Ile Ile Asn Ser Asn 385 390 395 400 Tyr Thr Asp Glu Glu Asp Asp Tyr Asp Ile Ala Leu Met Arg Leu Ser 405 410 415 Lys Pro Leu Thr Leu Ser Ala His Ile His Pro Ala Cys Leu Pro Met 420 425 430 His Gly Gln Thr Phe Ser Leu Asn Glu Thr Cys Trp Ile Thr Gly Phe 435 440 445 Gly Lys Thr Arg Glu Thr Asp Asp Lys Thr Ser Pro Phe Leu Arg Glu 450 455 460 Val Gln Val Asn Leu Ile Asp Phe Lys Lys Cys Asn Asp Tyr Leu Val 465 470 475 480 Tyr Asp Ser Tyr Leu Thr Pro Arg Met Met Cys Ala Gly Asp Leu Arg 485 490 495 Gly Gly Arg Asp Ser Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Cys 500 505 510 Glu Gln Asn Asn Arg Trp Tyr Leu Ala Gly Val Thr Ser Trp Gly Thr 515 520 525 Gly Cys Gly Gln Arg Asn Lys Pro Gly Val Tyr Thr Lys Val Thr Glu 530 535 540 Val Leu Pro Trp Ile Tyr Ser Lys Met Glu Asn Arg Ala Gln Arg Val 545 550 555 560 Glu Lys Ala Trp Thr Tyr Arg Pro Gly Arg Gln Leu Leu Gly Arg Cys 565 570 575 Ser Pro Arg Ser Ile Phe Leu Cys Lys Val Ala Met Asp Phe Glu Asn 580 585 590 Val Ser Val Ser Ala Glu Asp Phe Val Ile Val Phe Val Ile Lys His 595 600 605 Leu Cys Met Gly Ile Arg Ser Ser Trp Pro Phe Pro Ala Leu Phe Val 610 615 620 Leu Val Phe Leu Ile Phe Phe Leu Leu Leu Leu Leu Ser Phe Leu Lys 625 630 635 640 Asn Thr Ser Asp Ser Ile Leu Thr Leu Thr Thr Phe Thr Ala Val Thr 645 650 655 Arg Met Leu Pro Glu Asn Tyr His Ser Phe Pro Phe Pro Leu Lys Ile 660 665 670 Phe His Gln Asn Leu Thr Thr Ile Ile Lys Glu Tyr Lys Val Ile Lys 675 680 685 <210> SEQ ID NO 27 <211> LENGTH: 1471 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: misc_feature <222> LOCATION: (626)...(1324) <223> OTHER INFORMATION: DESC1 gene <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (56)...(1324) <223> OTHER INFORMATION: protease domain <400> SEQUENCE: 27 tgacttggat gtagacctcg accttcacag gactcttcat tgctggttgg caatg atg 58 Met 1 tat cgg cca gat gtg gtg agg gct agg aaa aga gtt tgt tgg gaa ccc 106Tyr Arg Pro Asp Val Val Arg Ala Arg Lys Arg Val Cys Trp Glu Pro 5 10 15 tgg gtt atc ggc ctc gtc ats ttc ata tcc ctg att gtc ctg gca gtg 154 Trp Val Ile Gly Leu Val Xaa Phe Ile Ser Leu Ile Val Leu Ala Val 20 25 30 tgc att gga stc act gtt cat tat gtg aga tat aat caa aag aag acc 202 Cys Ile Gly Xaa Thr Val His Tyr Val Arg Tyr Asn Gln Lys Lys Thr 35 40 45 tac aat tac tat agc aca ttg tca ttt aca act gac aaa cta tat gct 250 Tyr Asn Tyr Tyr Ser Thr Leu Ser Phe Thr Thr Asp Lys Leu Tyr Ala 50 55 60 65 gag ttt ggc aga gag gct tct aac aat ttt aca gaa atg agc cag aga 298 Glu Phe Gly Arg Glu Ala Ser Asn Asn Phe Thr Glu Met Ser Gln Arg 70 75 80 ctt gaa tca atg gtg aaa aat gca ttt tat aaa tct cca tta agg gaa 346 Leu Glu Ser Met Val Lys Asn Ala Phe Tyr Lys Ser Pro Leu Arg Glu 85 90 95 gaa ttt gtc aag tct cag gtt atc aag ttc agt caa cag aag cat gga 394Glu Phe Val Lys Ser Gln Val Ile Lys Phe Ser Gln Gln Lys His Gly 100 105 110 gtg ttg gct cat atg ctg ttg att tgt aga ttt cac tct act gag gat 442 Val Leu Ala His Met Leu Leu Ile Cys Arg Phe His Ser Thr Glu Asp 115 120 125 cct gaa act gta gat aaa att gtt caa ctt gtt tta cat gaa aag ctg 490Pro Glu Thr Val Asp Lys Ile Val Gln Leu Val Leu His Glu Lys Leu 130 135 140 145 caa gat gct gta gga ccc cct aaa gta gat cct cac tca gtt aaa att 538 Gln Asp Ala Val Gly Pro Pro Lys Val Asp Pro His Ser Val Lys Ile 150 155 160 aaa aaa atc aac aag aca gaa aca gac agc tat cta aac cat tgc tgc 586 Lys Lys Ile Asn Lys Thr Glu Thr Asp Ser Tyr Leu Asn His Cys Cys 165 170 175 gga aca cga aga agt aaa act cta ggt cag agt ctc agg atc gtt ggt 634 Gly Thr Arg Arg Ser Lys Thr Leu Gly Gln Ser Leu Arg Ile Val Gly 180 185 190 ggg aca gaa gta gaa gag ggt gaa tgg ccc tgg cag gct agc ctg cag 682 Gly Thr Glu Val Glu Glu Gly Glu Trp Pro Trp Gln Ala Ser Leu Gln 195 200 205 tgg gat ggg agt cat cgc tgt gga gca acc tta att aat gcc aca tgg 730 Trp Asp Gly Ser His Arg Cys Gly Ala Thr Leu Ile Asn Ala Thr Trp 210 215 220 225 ctt gtg agt gct gct cac tgt ttt aca aca tat aag aac cct gcc aga 778 Leu Val Ser Ala Ala His Cys Phe Thr Thr Tyr Lys Asn Pro Ala Arg 230 235 240 tgg act gct tcc ttt gga gta aca ata aaa cct tcg aaa atg aaa cgg 826 Trp Thr Ala Ser Phe Gly Val Thr Ile Lys Pro Ser Lys Met Lys Arg 245 250 255 ggt ctc cgg aga ata att gtc cat gaa aaa tac aaa cac cca tca cat 874 Gly Leu Arg Arg Ile Ile Val His Glu Lys Tyr Lys His Pro Ser His 260 265 270 gac tat gat att tct ctt gca gag ctt tct agc cct gtt ccc tac aca 922 Asp Tyr Asp Ile Ser Leu Ala Glu Leu Ser Ser Pro Val Pro Tyr Thr 275 280 285 aat gca gta cat aga gtt tgt ctc cct gat gca tcc tat gag ttt caa 970 Asn Ala Val His Arg Val Cys Leu Pro Asp Ala Ser Tyr Glu Phe Gln 290 295 300 305 cca ggt gat gtg atg ttt gtg aca gga ttt gga gca ctg aaa aat gat 1018 Pro Gly Asp Val Met Phe Val Thr Gly Phe Gly Ala Leu Lys Asn Asp 310 315 320 ggt tac agt caa aat cat ctt cga caa gca cag gtg act ctc ata gac 1066 Gly Tyr Ser Gln Asn His Leu Arg Gln Ala Gln Val Thr Leu Ile Asp 325 330 335 gct aca act tgc aat gaa cct caa gct tac aat gac gcc ata act cct 1114 Ala Thr Thr Cys Asn Glu Pro Gln Ala Tyr Asn Asp Ala Ile Thr Pro 340 345 350 aga atg tta tgt gct ggc tcc tta gaa gga aaa aca gat gca tgc cag 1162 Arg Met Leu Cys Ala Gly Ser Leu Glu Gly Lys Thr Asp Ala Cys Gln 355 360 365 ggt gac tct gga gga cca ctg gtt agt tca gat gct aga gat atc tgg 1210 Gly Asp Ser Gly Gly Pro Leu Val Ser Ser Asp Ala Arg Asp Ile Trp 370 375 380 385 tac ctt gct gga ata gtg agc tsg gga gat gaa tgt gcg aaa ccc aac 1258 Tyr Leu Ala Gly Ile Val Ser Xaa Gly Asp Glu Cys Ala Lys Pro Asn 390 395 400 aag cct ggt gtt tat act aga gtt acg gcc ttg cgg gac tgg att act 1306 Lys Pro Gly Val Tyr Thr Arg Val Thr Ala Leu Arg Asp Trp Ile Thr 405 410 415 tca aaa act ggt atc taa gagagaaaag cctcatggaa cagataacat 1354 Ser Lys Thr Gly Ile * 420 ttttttttgt tttttgggtg tggaggccat ttttagagat acagaattgg agaagacttg 1414 caaaacagct agatttgact gatctcaata aactgtttgc ttgatgcaaa aaaaaaa 1471 <210> SEQ ID NO 28 <211> LENGTH: 4933 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (94)...(3222) <223> OTHER INFORMATION: Nucleotide sequence encoding corin <300> PUBLICATION INFORMATION: <308> DATABASE ACCESSION NUMBER: GenBank AF133845 <309> DATABASE ENTRY DATE: 1999-05-24 <400> SEQUENCE: 28 aaatcatccg tagtgcctcc ccgggggaca cgtagaggag agaaaagcga ccaagataaa 60 agtggacaga agaataagcg agacttttta tcc atg aaa cag tct cct gcc ctc 114 Met Lys Gln Ser Pro Ala Leu 1 5 gct ccg gaa gag cgc tac cgc aga gcc ggg tcc cca aag ccg gtc ttg 162 Ala Pro Glu Glu Arg Tyr Arg Arg Ala Gly Ser Pro Lys Pro Val Leu 10 15 20 aga gct gat gac aat aac atg ggc aat ggc tgc tct cag aag ctg gcg 210 Arg Ala Asp Asp Asn Asn Met Gly Asn Gly Cys Ser Gln Lys Leu Ala 25 30 35 act gct aac ctc ctc cgg ttc cta ttg ctg gtc ctg att cca tgt atc 258 Thr Ala Asn Leu Leu Arg Phe Leu Leu Leu Val Leu Ile Pro Cys Ile 40 45 50 55 tgt gct ctc gtt ctc ttg ctg gtg atc ctg ctt tcc tat gtt gga aca 306 Cys Ala Leu Val Leu Leu Leu Val Ile Leu Leu Ser Tyr Val Gly Thr 60 65 70 tta caa aag gtc tat ttt aaa tca aat ggg agt gaa cct ttg gtc act 354 Leu Gln Lys Val Tyr Phe Lys Ser Asn Gly Ser Glu Pro Leu Val Thr 75 80 85 gat ggt gaa atc caa ggg tcc gat gtt att ctt aca aat aca att tat 402 Asp Gly Glu Ile Gln Gly Ser Asp Val Ile Leu Thr Asn Thr Ile Tyr 90 95 100 aac cag agc act gtg gtg tct act gca cat ccc gac caa cac gtt cca 450 Asn Gln Ser Thr Val Val Ser Thr Ala His Pro Asp Gln His Val Pro 105 110 115 gcc tgg act acg gat gct tct ctc cca ggg gac caa agt cac agg aat 498Ala Trp Thr Thr Asp Ala Ser Leu Pro Gly Asp Gln Ser His Arg Asn 120 125 130 135 aca agt gcc tgt atg aac atc acc cac agc cag tgt cag atg ctg ccc 546Thr Ser Ala Cys Met Asn Ile Thr His Ser Gln Cys Gln Met Leu Pro 140 145 150 tac cac gcc acg ctg aca cct ctc ctc tca gtt gtc aga aac atg gaa 594 Tyr His Ala Thr Leu Thr Pro Leu Leu Ser Val Val Arg Asn Met Glu 155 160 165 atg gaa aag ttc ctc aag ttt ttc aca tat ctc cat cgc ctc agt tgc 642 Met Glu Lys Phe Leu Lys Phe Phe Thr Tyr Leu His Arg Leu Ser Cys 170 175 180 tat caa cat atc atg ctg ttt ggc tgt acc ctc gcc ttc cct gag tgc 690 Tyr Gln His Ile Met Leu Phe Gly Cys Thr Leu Ala Phe Pro Glu Cys 185 190 195 atc att gat ggc gat gac agt cat gga ctc ctg ccc tgt agg tcc ttc 738 Ile Ile Asp Gly Asp Asp Ser His Gly Leu Leu Pro Cys Arg Ser Phe 200 205 210 215 tgt gag gct gca aaa gaa ggc tgt gaa tca gtc ctg ggg atg gtg aat 786 Cys Glu Ala Ala Lys Glu Gly Cys Glu Ser Val Leu Gly Met Val Asn 220 225 230 tac tcc tgg ccg gat ttc ctc aga tgc tcc cag ttt aga aac caa act 834 Tyr Ser Trp Pro Asp Phe Leu Arg Cys Ser Gln Phe Arg Asn Gln Thr 235 240 245 gaa agc agc aat gtc agc aga att tgc ttc tca cct cag cag gaa aac 882 Glu Ser Ser Asn Val Ser Arg Ile Cys Phe Ser Pro Gln Gln Glu Asn 250 255 260 gga aag caa ttg ctc tgt gga agg ggt gag aac ttt ctg tgt gcc agt 930 Gly Lys Gln Leu Leu Cys Gly Arg Gly Glu Asn Phe Leu Cys Ala Ser 265 270 275 gga atc tgc atc ccc ggg aaa ctg caa tgt aat ggc tac aac gac tgt 978 Gly Ile Cys Ile Pro Gly Lys Leu Gln Cys Asn Gly Tyr Asn Asp Cys 280 285 290 295 gac gac tgg agt gac gag gct cat tgc aac tgc agc gag aat ctg ttt 1026 Asp Asp Trp Ser Asp Glu Ala His Cys Asn Cys Ser Glu Asn Leu Phe 300 305 310 cac tgt cac aca ggc aag tgc ctt aat tac agc ctt gtg tgt gat gga 1074 His Cys His Thr Gly Lys Cys Leu Asn Tyr Ser Leu Val Cys Asp Gly 315 320 325 tat gat gac tgt ggg gat ttg agt gat gag caa aac tgt gat tgc aat 1122 Tyr Asp Asp Cys Gly Asp Leu Ser Asp Glu Gln Asn Cys Asp Cys Asn 330 335 340 ccc aca aca gag cat cgc tgc ggg gac ggg cgc tgc atc gcc atg gag 1170 Pro Thr Thr Glu His Arg Cys Gly Asp Gly Arg Cys Ile Ala Met Glu 345 350 355 tgg gtg tgt gat ggt gac cac gac tgt gtg gat aag tcc gac gag gtc 1218 Trp Val Cys Asp Gly Asp His Asp Cys Val Asp Lys Ser Asp Glu Val 360 365 370 375 aac tgc tcc tgt cac agc cag ggt ctg gtg gaa tgc aga aat gga caa 1266 Asn Cys Ser Cys His Ser Gln Gly Leu Val Glu Cys Arg Asn Gly Gln 380 385 390 tgt atc ccc agc acg ttt caa tgt gat ggt gac gag gac tgc aag gat 1314 Cys Ile Pro Ser Thr Phe Gln Cys Asp Gly Asp Glu Asp Cys Lys Asp 395 400 405 ggg agt gat gag gag aac tgc agc gtc att cag act tca tgt caa gaa 1362 Gly Ser Asp Glu Glu Asn Cys Ser Val Ile Gln Thr Ser Cys Gln Glu 410 415 420 gga gac caa aga tgc ctc tac aat ccc tgc ctt gat tca tgt ggt ggt 1410 Gly Asp Gln Arg Cys Leu Tyr Asn Pro Cys Leu Asp Ser Cys Gly Gly 425 430 435 agc tct ctc tgt gac ccg aac aac agt ctg aat aac tgt agt caa tgt 1458 Ser Ser Leu Cys Asp Pro Asn Asn Ser Leu Asn Asn Cys Ser Gln Cys 440 445 450 455 gaa cca att aca ttg gaa ctc tgc atg aat ttg ccc tac aac agt aca 1506 Glu Pro Ile Thr Leu Glu Leu Cys Met Asn Leu Pro Tyr Asn Ser Thr 460 465 470 agt tat cca aat tat ttt ggc cac agg act caa aag gaa gca tcc atc 1554 Ser Tyr Pro Asn Tyr Phe Gly His Arg Thr Gln Lys Glu Ala Ser Ile 475 480 485 agc tgg gag tct tct ctt ttc cct gca ctt gtt caa acc aac tgt tat 1602 Ser Trp Glu Ser Ser Leu Phe Pro Ala Leu Val Gln Thr Asn Cys Tyr 490 495 500 aaa tac ctc atg ttc ttt tct tgc acc att ttg gta cca aaa tgt gat 1650 Lys Tyr Leu Met Phe Phe Ser Cys Thr Ile Leu Val Pro Lys Cys Asp 505 510 515 gtg aat aca ggc gag cgt atc cct cct tgc agg gca ttg tgt gaa cac 1698 Val Asn Thr Gly Glu Arg Ile Pro Pro Cys Arg Ala Leu Cys Glu His 520 525 530 535 tct aaa gaa cgc tgt gag tct gtt ctt ggg att gtg ggc cta cag tgg 1746 Ser Lys Glu Arg Cys Glu Ser Val Leu Gly Ile Val Gly Leu Gln Trp 540 545 550 cct gaa gac aca gat tgc agt caa ttt cca gag gaa aat tca gac aat 1794 Pro Glu Asp Thr Asp Cys Ser Gln Phe Pro Glu Glu Asn Ser Asp Asn 555 560 565 caa acc tgc ctg atg cct gat gaa tat gtg gaa gaa tgc tca cct agt 1842 Gln Thr Cys Leu Met Pro Asp Glu Tyr Val Glu Glu Cys Ser Pro Ser 570 575 580 cat ttc aag tgc cgc tca gga cag tgt gtt ctg gct tcc aga aga tgt 1890 His Phe Lys Cys Arg Ser Gly Gln Cys Val Leu Ala Ser Arg Arg Cys 585 590 595 gat ggc cag gcc gac tgt gac gat gac agt gat gag gaa aac tgt ggt 1938 Asp Gly Gln Ala Asp Cys Asp Asp Asp Ser Asp Glu Glu Asn Cys Gly 600 605 610 615 tgt aaa gag aga gat ctt tgg gaa tgt cca tcc aat aaa caa tgt ttg 1986 Cys Lys Glu Arg Asp Leu Trp Glu Cys Pro Ser Asn Lys Gln Cys Leu 620 625 630 aag cac aca gtg atc tgc gat ggg ttc cca gac tgc cct gat tac atg 2034 Lys His Thr Val Ile Cys Asp Gly Phe Pro Asp Cys Pro Asp Tyr Met 635 640 645 gac gag aaa aac tgc tca ttt tgc caa gat gat gag ctg gaa tgt gca 2082 Asp Glu Lys Asn Cys Ser Phe Cys Gln Asp Asp Glu Leu Glu Cys Ala 650 655 660 aac cat gcg tgt gtg tca cgt gac ctg tgg tgt gat ggt gaa gcc gac 2130 Asn His Ala Cys Val Ser Arg Asp Leu Trp Cys Asp Gly Glu Ala Asp 665 670 675 tgc tca gac agt tca gat gaa tgg gac tgt gtg acc ctc tct ata aat 2178 Cys Ser Asp Ser Ser Asp Glu Trp Asp Cys Val Thr Leu Ser Ile Asn 680 685 690 695 gtg aac tcc tct tcc ttt ctg atg gtt cac aga gct gcc aca gaa cac 2226 Val Asn Ser Ser Ser Phe Leu Met Val His Arg Ala Ala Thr Glu His 700 705 710 cat gtg tgt gca gat ggc tgg cag gag ata ttg agt cag ctg gcc tgc 2274 His Val Cys Ala Asp Gly Trp Gln Glu Ile Leu Ser Gln Leu Ala Cys 715 720 725 aag cag atg ggt tta gga gaa cca tct gtg acc aaa ttg ata cag gaa 2322 Lys Gln Met Gly Leu Gly Glu Pro Ser Val Thr Lys Leu Ile Gln Glu 730 735 740 cag gag aaa gag ccg cgg tgg ctg aca tta cac tcc aac tgg gag agc 2370 Gln Glu Lys Glu Pro Arg Trp Leu Thr Leu His Ser Asn Trp Glu Ser 745 750 755 ctc aat ggg acc act tta cat gaa ctt cta gta aat ggg cag tct tgt 2418 Leu Asn Gly Thr Thr Leu His Glu Leu Leu Val Asn Gly Gln Ser Cys 760 765 770 775 gag agc aga agt aaa att tct ctt ctg tgt act aaa caa gac tgt ggg 2466 Glu Ser Arg Ser Lys Ile Ser Leu Leu Cys Thr Lys Gln Asp Cys Gly 780 785 790 cgc cgc cct gct gcc cga atg aac aaa agg atc ctt gga ggt cgg acg 2514 Arg Arg Pro Ala Ala Arg Met Asn Lys Arg Ile Leu Gly Gly Arg Thr 795 800 805 agt cgc cct gga agg tgg cca tgg cag tgt tct ctg cag agt gaa ccc 2562 Ser Arg Pro Gly Arg Trp Pro Trp Gln Cys Ser Leu Gln Ser Glu Pro 810 815 820 agt gga cat atc tgt ggc tgt gtc ctc att gcc aag aag tgg gtt ctg 2610 Ser Gly His Ile Cys Gly Cys Val Leu Ile Ala Lys Lys Trp Val Leu 825 830 835 aca gtt gcc cac tgc ttc gag ggg aga gag aat gct gca gtt tgg aaa 2658 Thr Val Ala His Cys Phe Glu Gly Arg Glu Asn Ala Ala Val Trp Lys 840 845 850 855 gtg gtg ctt ggc atc aac aat cta gac cat cca tca gtg ttc atg cag 2706 Val Val Leu Gly Ile Asn Asn Leu Asp His Pro Ser Val Phe Met Gln 860 865 870 aca cgc ttt gtg aag acc atc atc ctg cat ccc cgc tac agt cga gca 2754 Thr Arg Phe Val Lys Thr Ile Ile Leu His Pro Arg Tyr Ser Arg Ala 875 880 885 gtg gtg gac tat gac atc agc atc gtt gag ctg agt gaa gac atc agt 2802 Val Val Asp Tyr Asp Ile Ser Ile Val Glu Leu Ser Glu Asp Ile Ser 890 895 900 gag act ggc tac gtc cgg cct gtc tgc ttg ccc aac ccg gag cag tgg 2850 Glu Thr Gly Tyr Val Arg Pro Val Cys Leu Pro Asn Pro Glu Gln Trp 905 910 915 cta gag cct gac acg tac tgc tat atc aca ggc tgg ggc cac atg ggc 2898 Leu Glu Pro Asp Thr Tyr Cys Tyr Ile Thr Gly Trp Gly His Met Gly 920 925 930 935 aat aaa atg cca ttt aag ctg caa gag gga gag gtc cgc att att tct 2946 Asn Lys Met Pro Phe Lys Leu Gln Glu Gly Glu Val Arg Ile Ile Ser 940 945 950 ctg gaa cat tgt cag tcc tac ttt gac atg aag acc atc acc act cgg 2994 Leu Glu His Cys Gln Ser Tyr Phe Asp Met Lys Thr Ile Thr Thr Arg 955 960 965 atg ata tgt gct ggc tat gag tct ggc aca gtt gat tca tgc atg ggt 3042 Met Ile Cys Ala Gly Tyr Glu Ser Gly Thr Val Asp Ser Cys Met Gly 970 975 980 gac agc ggt ggg cct ctt gtt tgt gag aag cct gga gga cgg tgg aca 3090 Asp Ser Gly Gly Pro Leu Val Cys Glu Lys Pro Gly Gly Arg Trp Thr 985 990 995 tta ttt gga tta act tca tgg ggc tcc gtc tgc ttt tcc aaa gtc ctg 3138 Leu Phe Gly Leu Thr Ser Trp Gly Ser Val Cys Phe Ser Lys Val Leu 1000 1005 1010 1015 ggg cct ggc gtt tat agt aat gtg tca tat ttc gtc gaa tgg att aaa 3186 Gly Pro Gly Val Tyr Ser Asn Val Ser Tyr Phe Val Glu Trp Ile Lys 1020 1025 1030 aga cag att tac atc cag acc ttt ctc cta aac taa ttataaggat 3232 Arg Gln Ile Tyr Ile Gln Thr Phe Leu Leu Asn * 1035 1040 gatcagagac ttttgccagc tacactaaaa gaaaatggcc ttcttgactg tgaagagctg 3292 cctgcagaga gctgtacaga agcacttttc atggacagaa atgctcaatc gtgcactgca 3352 aatttgcatg tttgttttgg actaattttt ttcaatttat tttttcacct tcatttttct 3412 cttatttcaa gttcaatgaa agactttaca aaagcaaaca aagcagactt tgtccttttg 3472 ccaggcctaa ccatgactgc agcacaaaat tatcgactct ggcgagattt aaaatcaggt 3532 gctacagtaa caggttatgg aatggtctct tttatcctat cacaaaaaaa gacatagata 3592 tttaggctga ttaattatct ctaccagttt ttgtttctca agctcagtgc atagtggtaa 3652 atttcagtgt taacattgga gacttgcttt tctttttctt tttttatacc ccacaattct 3712 tttttattac acttcgaatt ttagggtaca cgagcacaac gtgcaggtta gttacatatg 3772 tatacatgtg ccatgttggt gtgctgaacc cagtaactcg tcatttgatt tattaaaagc 3832 caagataatt tacatgttta aagtatttac tattaccccc ttctaatgtt tgcataattc 3892 tgagaactga taaaagacag caataaaaga ccagtgtcat ccatttaggt agcaagacat 3952 attgaatgca aagttcttta gatatcaata ttaacacttg acattattgg accccccatt 4012 ctggatgtat atcaagatca taattttata gaagagtctc tatagaactg tcctcatagc 4072 tgggtttgtt caggatatat gagttggctg attgagactg caacaactac atctatattt 4132 atgggcaata ttttgtttta cttatgtggc aaagaactgg atattaaact ttgcaaaaga 4192 gaatttagat gagagatgca attttttaaa aagaaaatta atttgcatcc ctcgtttaat 4252 taaatttatt tttcagtttt cttgcgttca tccataccaa caaagtcata aagagcatat 4312 tttagagcac agtaagactt tgcatggagt aaaacatttt gtaattttcc tcaaaagatg 4372 tttaatatct ggtttcttct cattggtaat taaaatttta gaaatgattt ttagctctag 4432 gccactttac gcaactcaat ttctgaagca attagtggta aaaagtattt ttccccacta 4492 aaaaacttta aaacacaaat cttcatatat acttaattta attagtcagg catccatttt 4552 gccttttaaa caactaggat tccctactaa cctccaccag caacctggac tgcctcagca 4612 ttccaaatag atactacctg caattttata catgtatttt tgtatctttt ctgtgtgtaa 4672 acatagttga aattcaaaaa gttgtagcaa tttctatact attcatctcc tgtccttcag 4732 tttgtataaa cctaaggaga gtgtgaaatc cagcaactga attgtggtca cgattgtatg 4792 aaagttcaag aacatatgtc agttttgtta cagttgtagc tacatactca atgtatcaac 4852 ttttagcctg ctcaacttag gctcagtgaa atatatatat tatacttatt ttaaataatt 4912 cttaatacaa ataaaatggt a 4933 <210> SEQ ID NO 29 <211> LENGTH: 1042 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 29 Met Lys Gln Ser Pro Ala Leu Ala Pro Glu Glu Arg Tyr Arg Arg Ala 1 5 10 15 Gly Ser Pro Lys Pro Val Leu Arg Ala Asp Asp Asn Asn Met Gly Asn 20 25 30 Gly Cys Ser Gln Lys Leu Ala Thr Ala Asn Leu Leu Arg Phe Leu Leu 35 40 45 Leu Val Leu Ile Pro Cys Ile Cys Ala Leu Val Leu Leu Leu Val Ile 50 55 60 Leu Leu Ser Tyr Val Gly Thr Leu Gln Lys Val Tyr Phe Lys Ser Asn 65 70 75 80 Gly Ser Glu Pro Leu Val Thr Asp Gly Glu Ile Gln Gly Ser Asp Val 85 90 95 Ile Leu Thr Asn Thr Ile Tyr Asn Gln Ser Thr Val Val Ser Thr Ala 100 105 110 His Pro Asp Gln His Val Pro Ala Trp Thr Thr Asp Ala Ser Leu Pro 115 120 125 Gly Asp Gln Ser His Arg Asn Thr Ser Ala Cys Met Asn Ile Thr His 130 135 140 Ser Gln Cys Gln Met Leu Pro Tyr His Ala Thr Leu Thr Pro Leu Leu 145 150 155 160 Ser Val Val Arg Asn Met Glu Met Glu Lys Phe Leu Lys Phe Phe Thr 165 170 175 Tyr Leu His Arg Leu Ser Cys Tyr Gln His Ile Met Leu Phe Gly Cys 180 185 190 Thr Leu Ala Phe Pro Glu Cys Ile Ile Asp Gly Asp Asp Ser His Gly 195 200 205 Leu Leu Pro Cys Arg Ser Phe Cys Glu Ala Ala Lys Glu Gly Cys Glu 210 215 220 Ser Val Leu Gly Met Val Asn Tyr Ser Trp Pro Asp Phe Leu Arg Cys 225 230 235 240 Ser Gln Phe Arg Asn Gln Thr Glu Ser Ser Asn Val Ser Arg Ile Cys 245 250 255 Phe Ser Pro Gln Gln Glu Asn Gly Lys Gln Leu Leu Cys Gly Arg Gly 260 265 270 Glu Asn Phe Leu Cys Ala Ser Gly Ile Cys Ile Pro Gly Lys Leu Gln 275 280 285 Cys Asn Gly Tyr Asn Asp Cys Asp Asp Trp Ser Asp Glu Ala His Cys 290 295 300 Asn Cys Ser Glu Asn Leu Phe His Cys His Thr Gly Lys Cys Leu Asn 305 310 315 320 Tyr Ser Leu Val Cys Asp Gly Tyr Asp Asp Cys Gly Asp Leu Ser Asp 325 330 335 Glu Gln Asn Cys Asp Cys Asn Pro Thr Thr Glu His Arg Cys Gly Asp 340 345 350 Gly Arg Cys Ile Ala Met Glu Trp Val Cys Asp Gly Asp His Asp Cys 355 360 365 Val Asp Lys Ser Asp Glu Val Asn Cys Ser Cys His Ser Gln Gly Leu 370 375 380 Val Glu Cys Arg Asn Gly Gln Cys Ile Pro Ser Thr Phe Gln Cys Asp 385 390 395 400 Gly Asp Glu Asp Cys Lys Asp Gly Ser Asp Glu Glu Asn Cys Ser Val 405 410 415 Ile Gln Thr Ser Cys Gln Glu Gly Asp Gln Arg Cys Leu Tyr Asn Pro 420 425 430 Cys Leu Asp Ser Cys Gly Gly Ser Ser Leu Cys Asp Pro Asn Asn Ser 435 440 445 Leu Asn Asn Cys Ser Gln Cys Glu Pro Ile Thr Leu Glu Leu Cys Met 450 455 460 Asn Leu Pro Tyr Asn Ser Thr Ser Tyr Pro Asn Tyr Phe Gly His Arg 465 470 475 480 Thr Gln Lys Glu Ala Ser Ile Ser Trp Glu Ser Ser Leu Phe Pro Ala 485 490 495 Leu Val Gln Thr Asn Cys Tyr Lys Tyr Leu Met Phe Phe Ser Cys Thr 500 505 510 Ile Leu Val Pro Lys Cys Asp Val Asn Thr Gly Glu Arg Ile Pro Pro 515 520 525 Cys Arg Ala Leu Cys Glu His Ser Lys Glu Arg Cys Glu Ser Val Leu 530 535 540 Gly Ile Val Gly Leu Gln Trp Pro Glu Asp Thr Asp Cys Ser Gln Phe 545 550 555 560 Pro Glu Glu Asn Ser Asp Asn Gln Thr Cys Leu Met Pro Asp Glu Tyr 565 570 575 Val Glu Glu Cys Ser Pro Ser His Phe Lys Cys Arg Ser Gly Gln Cys 580 585 590 Val Leu Ala Ser Arg Arg Cys Asp Gly Gln Ala Asp Cys Asp Asp Asp 595 600 605 Ser Asp Glu Glu Asn Cys Gly Cys Lys Glu Arg Asp Leu Trp Glu Cys 610 615 620 Pro Ser Asn Lys Gln Cys Leu Lys His Thr Val Ile Cys Asp Gly Phe 625 630 635 640 Pro Asp Cys Pro Asp Tyr Met Asp Glu Lys Asn Cys Ser Phe Cys Gln 645 650 655 Asp Asp Glu Leu Glu Cys Ala Asn His Ala Cys Val Ser Arg Asp Leu 660 665 670 Trp Cys Asp Gly Glu Ala Asp Cys Ser Asp Ser Ser Asp Glu Trp Asp 675 680 685 Cys Val Thr Leu Ser Ile Asn Val Asn Ser Ser Ser Phe Leu Met Val 690 695 700 His Arg Ala Ala Thr Glu His His Val Cys Ala Asp Gly Trp Gln Glu 705 710 715 720 Ile Leu Ser Gln Leu Ala Cys Lys Gln Met Gly Leu Gly Glu Pro Ser 725 730 735 Val Thr Lys Leu Ile Gln Glu Gln Glu Lys Glu Pro Arg Trp Leu Thr 740 745 750 Leu His Ser Asn Trp Glu Ser Leu Asn Gly Thr Thr Leu His Glu Leu 755 760 765 Leu Val Asn Gly Gln Ser Cys Glu Ser Arg Ser Lys Ile Ser Leu Leu 770 775 780 Cys Thr Lys Gln Asp Cys Gly Arg Arg Pro Ala Ala Arg Met Asn Lys 785 790 795 800 Arg Ile Leu Gly Gly Arg Thr Ser Arg Pro Gly Arg Trp Pro Trp Gln 805 810 815 Cys Ser Leu Gln Ser Glu Pro Ser Gly His Ile Cys Gly Cys Val Leu 820 825 830 Ile Ala Lys Lys Trp Val Leu Thr Val Ala His Cys Phe Glu Gly Arg 835 840 845 Glu Asn Ala Ala Val Trp Lys Val Val Leu Gly Ile Asn Asn Leu Asp 850 855 860 His Pro Ser Val Phe Met Gln Thr Arg Phe Val Lys Thr Ile Ile Leu 865 870 875 880 His Pro Arg Tyr Ser Arg Ala Val Val Asp Tyr Asp Ile Ser Ile Val 885 890 895 Glu Leu Ser Glu Asp Ile Ser Glu Thr Gly Tyr Val Arg Pro Val Cys 900 905 910 Leu Pro Asn Pro Glu Gln Trp Leu Glu Pro Asp Thr Tyr Cys Tyr Ile 915 920 925 Thr Gly Trp Gly His Met Gly Asn Lys Met Pro Phe Lys Leu Gln Glu 930 935 940 Gly Glu Val Arg Ile Ile Ser Leu Glu His Cys Gln Ser Tyr Phe Asp 945 950 955 960 Met Lys Thr Ile Thr Thr Arg Met Ile Cys Ala Gly Tyr Glu Ser Gly 965 970 975 Thr Val Asp Ser Cys Met Gly Asp Ser Gly Gly Pro Leu Val Cys Glu 980 985 990 Lys Pro Gly Gly Arg Trp Thr Leu Phe Gly Leu Thr Ser Trp Gly Ser 995 1000 1005 Val Cys Phe Ser Lys Val Leu Gly Pro Gly Val Tyr Ser Asn Val Ser 1010 1015 1020 Tyr Phe Val Glu Trp Ile Lys Arg Gln Ile Tyr Ile Gln Thr Phe Leu 1025 1030 1035 1040 Leu Asn <210> SEQ ID NO 30 <211> LENGTH: 3696 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (41)...(3100) <223> OTHER INFORMATION: Nucleic acid encoding human enterokinase <300> PUBLICATION INFORMATION: <308> DATABASE ACCESSION NUMBER: GenBank HSU09860 <309> DATABASE ENTRY DATE: 1995-06-03 <400> SEQUENCE: 30 accagacagt tcttaaatta gcaagccttc aaaaccaaaa atg ggg tcg aaa aga 55 Met Gly Ser Lys Arg 1 5 ggc ata tct tct agg cat cat tct ctc agc tcc tat gaa atc atg ttt 103 Gly Ile Ser Ser Arg His His Ser Leu Ser Ser Tyr Glu Ile Met Phe 10 15 20 gca gct ctc ttt gcc ata ttg gta gtg ctc tgt gct gga tta att gca 151 Ala Ala Leu Phe Ala Ile Leu Val Val Leu Cys Ala Gly Leu Ile Ala 25 30 35 gta tcc tgc ctg aca atc aag gaa tcc caa cga ggt gca gca ctt gga 199 Val Ser Cys Leu Thr Ile Lys Glu Ser Gln Arg Gly Ala Ala Leu Gly 40 45 50 cag agt cat gaa gcc aga gcg aca ttt aaa ata aca tcc gga gtt aca 247 Gln Ser His Glu Ala Arg Ala Thr Phe Lys Ile Thr Ser Gly Val Thr 55 60 65 tat aat cct aat ttg caa gac aaa ctc tca gtg gat ttc aaa gtt ctt 295Tyr Asn Pro Asn Leu Gln Asp Lys Leu Ser Val Asp Phe Lys Val Leu 70 75 80 85 gct ttt gac ctt cag caa atg ata gat gag atc ttt cta tca agc aat 343Ala Phe Asp Leu Gln Gln Met Ile Asp Glu Ile Phe Leu Ser Ser Asn 90 95 100 ctg aag aat gaa tat aag aac tca aga gtt tta caa ttt gaa aat ggc 391Leu Lys Asn Glu Tyr Lys Asn Ser Arg Val Leu Gln Phe Glu Asn Gly 105 110 115 agc att ata gtc gta ttt gac ctt ttc ttt gcc cag tgg gtg tca gat 439 Ser Ile Ile Val Val Phe Asp Leu Phe Phe Ala Gln Trp Val Ser Asp 120 125 130 caa aat gta aaa gaa gaa ctg att caa ggc ctt gaa gca aat aaa tcc 487Gln Asn Val Lys Glu Glu Leu Ile Gln Gly Leu Glu Ala Asn Lys Ser 135 140 145 agc caa ctg gtc act ttc cat att gat ttg aac agc gtt gat atc cta 535Ser Gln Leu Val Thr Phe His Ile Asp Leu Asn Ser Val Asp Ile Leu 150 155 160 165 gac aag cta aca acc acc agt cat ctg gca act cca gga aat gtc tca 583 Asp Lys Leu Thr Thr Thr Ser His Leu Ala Thr Pro Gly Asn Val Ser 170 175 180 ata gag tgc ctg cct ggt tca agt cct tgt act gat gct cta acg tgt 631 Ile Glu Cys Leu Pro Gly Ser Ser Pro Cys Thr Asp Ala Leu Thr Cys 185 190 195 ata aaa gct gat tta ttt tgt gat gga gaa gta aac tgt cca gat ggt 679 Ile Lys Ala Asp Leu Phe Cys Asp Gly Glu Val Asn Cys Pro Asp Gly 200 205 210 tct gac gaa gac aat aaa atg tgt gcc aca gtt tgt gat gga aga ttt 727 Ser Asp Glu Asp Asn Lys Met Cys Ala Thr Val Cys Asp Gly Arg Phe 215 220 225 ttg tta act gga tca tct ggg tct ttc cag gct act cat tat cca aaa 775 Leu Leu Thr Gly Ser Ser Gly Ser Phe Gln Ala Thr His Tyr Pro Lys 230 235 240 245 cct tct gaa aca agt gtt gtc tgc cag tgg atc ata cgt gta aac caa 823 Pro Ser Glu Thr Ser Val Val Cys Gln Trp Ile Ile Arg Val Asn Gln 250 255 260 gga ctt tcc att aaa ctg agc ttc gat gat ttt aat aca tat tat aca 871 Gly Leu Ser Ile Lys Leu Ser Phe Asp Asp Phe Asn Thr Tyr Tyr Thr 265 270 275 gat ata tta gat att tat gaa ggt gta gga tca agc aag att tta aga 919 Asp Ile Leu Asp Ile Tyr Glu Gly Val Gly Ser Ser Lys Ile Leu Arg 280 285 290 gct tct att tgg gaa act aat cct ggc aca ata aga att ttt tcc aac 967 Ala Ser Ile Trp Glu Thr Asn Pro Gly Thr Ile Arg Ile Phe Ser Asn 295 300 305 caa gtt act gcc acc ttt ctt ata gaa tct gat gaa agt gat tat gtt 1015 Gln Val Thr Ala Thr Phe Leu Ile Glu Ser Asp Glu Ser Asp Tyr Val 310 315 320 325 ggc ttt aat gca aca tat act gca ttt aac agc agt gag ctt aat aat 1063 Gly Phe Asn Ala Thr Tyr Thr Ala Phe Asn Ser Ser Glu Leu Asn Asn 330 335 340 tat gag aaa att aat tgt aac ttt gag gat ggc ttt tgt ttc tgg gtc 1111 Tyr Glu Lys Ile Asn Cys Asn Phe Glu Asp Gly Phe Cys Phe Trp Val 345 350 355 cag gat cta aat gat gat aat gaa tgg gaa agg att cag gga agc acc 1159 Gln Asp Leu Asn Asp Asp Asn Glu Trp Glu Arg Ile Gln Gly Ser Thr 360 365 370 ttt tct cct ttt act gga ccc aat ttt gac cac act ttt ggc aat gct 1207 Phe Ser Pro Phe Thr Gly Pro Asn Phe Asp His Thr Phe Gly Asn Ala 375 380 385 tca gga ttt tac att tct acc cca act gga cca gga ggg aga caa gaa 1255 Ser Gly Phe Tyr Ile Ser Thr Pro Thr Gly Pro Gly Gly Arg Gln Glu 390 395 400 405 cga gtg ggg ctt tta agc ctc cct ttg gac ccc act ttg gag cca gct 1303 Arg Val Gly Leu Leu Ser Leu Pro Leu Asp Pro Thr Leu Glu Pro Ala 410 415 420 tgc ctt agt ttc tgg tat cat atg tat ggt gaa aat gtc cat aaa tta 1351 Cys Leu Ser Phe Trp Tyr His Met Tyr Gly Glu Asn Val His Lys Leu 425 430 435 agc att aat atc agc aat gac caa aat atg gag aag aca gtt ttc caa 1399 Ser Ile Asn Ile Ser Asn Asp Gln Asn Met Glu Lys Thr Val Phe Gln 440 445 450 aag gaa gga aat tat gga gac aat tgg aat tat gga caa gta acc cta 1447 Lys Glu Gly Asn Tyr Gly Asp Asn Trp Asn Tyr Gly Gln Val Thr Leu 455 460 465 aat gaa aca gtt aaa ttt aag gtt gct ttt aat gct ttt aaa aac aag 1495 Asn Glu Thr Val Lys Phe Lys Val Ala Phe Asn Ala Phe Lys Asn Lys 470 475 480 485 atc ctg agt gat att gcg ttg gat gac att agc cta aca tat ggg att 1543 Ile Leu Ser Asp Ile Ala Leu Asp Asp Ile Ser Leu Thr Tyr Gly Ile 490 495 500 tgc aat ggg agt ctt tat cca gaa cca act ttg gtg cca act cct cca 1591 Cys Asn Gly Ser Leu Tyr Pro Glu Pro Thr Leu Val Pro Thr Pro Pro 505 510 515 cca gaa ctt cct acg gac tgt gga gga cct ttt gag ctg tgg gag cca 1639 Pro Glu Leu Pro Thr Asp Cys Gly Gly Pro Phe Glu Leu Trp Glu Pro 520 525 530 aat aca aca ttc agt tct acg aac ttt cca aac agc tac cct aat ctg 1687 Asn Thr Thr Phe Ser Ser Thr Asn Phe Pro Asn Ser Tyr Pro Asn Leu 535 540 545 gct ttc tgt gtt tgg att tta aat gca caa aaa gga aag aat ata caa 1735 Ala Phe Cys Val Trp Ile Leu Asn Ala Gln Lys Gly Lys Asn Ile Gln 550 555 560 565 ctt cat ttt caa gaa ttt gac tta gaa aat att aac gat gta gtt gaa 1783 Leu His Phe Gln Glu Phe Asp Leu Glu Asn Ile Asn Asp Val Val Glu 570 575 580 ata aga gat ggt gaa gaa gct gat tcc ttg ctc tta gct gtg tac aca 1831 Ile Arg Asp Gly Glu Glu Ala Asp Ser Leu Leu Leu Ala Val Tyr Thr 585 590 595 ggg cct ggc cca gta aag gat gtg ttc tct acc acc aac aga atg act 1879 Gly Pro Gly Pro Val Lys Asp Val Phe Ser Thr Thr Asn Arg Met Thr 600 605 610 gtg ctt ctc atc act aac gat gtg ttg gca aga gga ggg ttt aaa gca 1927 Val Leu Leu Ile Thr Asn Asp Val Leu Ala Arg Gly Gly Phe Lys Ala 615 620 625 aac ttt act act ggc tat cac ttg ggg att cca gag cca tgc aag gca 1975 Asn Phe Thr Thr Gly Tyr His Leu Gly Ile Pro Glu Pro Cys Lys Ala 630 635 640 645 gac cat ttt caa tgt aaa aat gga gag tgt gtt cca ctg gtg aat ctc 2023 Asp His Phe Gln Cys Lys Asn Gly Glu Cys Val Pro Leu Val Asn Leu 650 655 660 tgt gac ggt cat ctg cac tgt gag gat ggc tca gat gaa gca gat tgt 2071 Cys Asp Gly His Leu His Cys Glu Asp Gly Ser Asp Glu Ala Asp Cys 665 670 675 gtg cgt ttt ttc aat ggc aca acg aac aac aat ggt tta gtg cgg ttc 2119 Val Arg Phe Phe Asn Gly Thr Thr Asn Asn Asn Gly Leu Val Arg Phe 680 685 690 aga atc cag agc ata tgg cat aca gct tgt gct gag aac tgg acc acc 2167 Arg Ile Gln Ser Ile Trp His Thr Ala Cys Ala Glu Asn Trp Thr Thr 695 700 705 cag att tca aat gat gtt tgt caa ctg ctg gga cta ggg agt gga aac 2215 Gln Ile Ser Asn Asp Val Cys Gln Leu Leu Gly Leu Gly Ser Gly Asn 710 715 720 725 tca tca aag cca atc ttc tct acc gat ggt gga cca ttt gtc aaa tta 2263 Ser Ser Lys Pro Ile Phe Ser Thr Asp Gly Gly Pro Phe Val Lys Leu 730 735 740 aac aca gca cct gat ggc cac tta ata cta aca ccc agt caa cag tgt 2311 Asn Thr Ala Pro Asp Gly His Leu Ile Leu Thr Pro Ser Gln Gln Cys 745 750 755 tta cag gat tcc ttg att cgg tta cag tgt aac cat aaa tct tgt gga 2359 Leu Gln Asp Ser Leu Ile Arg Leu Gln Cys Asn His Lys Ser Cys Gly 760 765 770 aaa aaa ctg gca gct caa gac atc acc cca aag att gtt gga gga agt 2407 Lys Lys Leu Ala Ala Gln Asp Ile Thr Pro Lys Ile Val Gly Gly Ser 775 780 785 aat gcc aaa gaa ggg gcc tgg ccc tgg gtt gtg ggt ctg tat tat ggc 2455 Asn Ala Lys Glu Gly Ala Trp Pro Trp Val Val Gly Leu Tyr Tyr Gly 790 795 800 805 ggc cga ctg ctc tgc ggc gca tct ctc gtc agc agt gac tgg ctg gtg 2503 Gly Arg Leu Leu Cys Gly Ala Ser Leu Val Ser Ser Asp Trp Leu Val 810 815 820 tcc gcc gca cac tgc gtg tat ggg aga aac tta gag cca tcc aag tgg 2551 Ser Ala Ala His Cys Val Tyr Gly Arg Asn Leu Glu Pro Ser Lys Trp 825 830 835 aca gca atc cta ggc ctg cat atg aaa tca aat ctg acc tct cct caa 2599 Thr Ala Ile Leu Gly Leu His Met Lys Ser Asn Leu Thr Ser Pro Gln 840 845 850 aca gtc cct cga tta ata gat gaa att gtc ata aac cct cat tac aat 2647 Thr Val Pro Arg Leu Ile Asp Glu Ile Val Ile Asn Pro His Tyr Asn 855 860 865 agg cga aga aag gac aac gac att gcc atg atg cat ctg gaa ttt aaa 2695 Arg Arg Arg Lys Asp Asn Asp Ile Ala Met Met His Leu Glu Phe Lys 870 875 880 885 gtg aat tac aca gat tac ata caa cct att tgt tta ccg gaa gaa aat 2743 Val Asn Tyr Thr Asp Tyr Ile Gln Pro Ile Cys Leu Pro Glu Glu Asn 890 895 900 caa gtt ttt cct cca gga aga aat tgt tct att gct ggt tgg ggg acg 2791 Gln Val Phe Pro Pro Gly Arg Asn Cys Ser Ile Ala Gly Trp Gly Thr 905 910 915 gtt gta tat caa ggt act act gca aac ata ttg caa gaa gct gat gtt 2839 Val Val Tyr Gln Gly Thr Thr Ala Asn Ile Leu Gln Glu Ala Asp Val 920 925 930 cct ctt cta tca aat gag aga tgc caa cag cag atg cca gaa tat aac 2887 Pro Leu Leu Ser Asn Glu Arg Cys Gln Gln Gln Met Pro Glu Tyr Asn 935 940 945 att act gaa aat atg ata tgt gca ggc tat gaa gaa gga gga ata gat 2935 Ile Thr Glu Asn Met Ile Cys Ala Gly Tyr Glu Glu Gly Gly Ile Asp 950 955 960 965 tct tgt cag ggg gat tca gga gga cca tta atg tgc caa gaa aac aac 2983 Ser Cys Gln Gly Asp Ser Gly Gly Pro Leu Met Cys Gln Glu Asn Asn 970 975 980 agg tgg ttc ctt gct ggt gtg acc tca ttt gga tac aag tgt gcc ctg 3031 Arg Trp Phe Leu Ala Gly Val Thr Ser Phe Gly Tyr Lys Cys Ala Leu 985 990 995 cct aat cgc ccc gga gtg tat gcc agg gtc tca agg ttt acc gaa tgg 3079 Pro Asn Arg Pro Gly Val Tyr Ala Arg Val Ser Arg Phe Thr Glu Trp 1000 1005 1010 ata caa agt ttt cta cat tag cgcatttctt aaactaaaca ggaaagtcgc 3130 Ile Gln Ser Phe Leu His * 1015 attattttcc cattctactc tagaaagcat ggaaattaag tgtttcgtac aaaaatttta 3190 aaaagttacc aaaggttttt attcttacct atgtcaatga aatgctaggg ggccagggaa 3250 acaaaatttt aaaaataata aaattcacca tagcaataca gaataacttt aaaataccat 3310 taaatacatt tgtatttcat tgtgaacagg tatttcttca cagatctcat ttttaaaatt 3370 cttaatgatt atttttatta cttactgttg tttaaaggga tgttatttta aagcatatac 3430 catacactta agaaatttga gcagaattta aaaaagaaag aaaataaatt gtttttccca 3490 aagtatgtca ctgttggaaa taaactgcca taaattttct agttccagtt tagtttgctg 3550 ctattagcag aaactcaatt gtttctctgt cttttctatc aaaattttca acatatgcat 3610 aaccttagta ttttcccaac caatagaaac tatttattgt aagcttatgt cacaggcctg 3670 gactaaattg attttacgtt cctctt 3696 <210> SEQ ID NO 31 <211> LENGTH: 1019 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 31 Met Gly Ser Lys Arg Gly Ile Ser Ser Arg His His Ser Leu Ser Ser 1 5 10 15 Tyr Glu Ile Met Phe Ala Ala Leu Phe Ala Ile Leu Val Val Leu Cys 20 25 30 Ala Gly Leu Ile Ala Val Ser Cys Leu Thr Ile Lys Glu Ser Gln Arg 35 40 45 Gly Ala Ala Leu Gly Gln Ser His Glu Ala Arg Ala Thr Phe Lys Ile 50 55 60 Thr Ser Gly Val Thr Tyr Asn Pro Asn Leu Gln Asp Lys Leu Ser Val 65 70 75 80 Asp Phe Lys Val Leu Ala Phe Asp Leu Gln Gln Met Ile Asp Glu Ile 85 90 95 Phe Leu Ser Ser Asn Leu Lys Asn Glu Tyr Lys Asn Ser Arg Val Leu 100 105 110 Gln Phe Glu Asn Gly Ser Ile Ile Val Val Phe Asp Leu Phe Phe Ala 115 120 125 Gln Trp Val Ser Asp Gln Asn Val Lys Glu Glu Leu Ile Gln Gly Leu 130 135 140 Glu Ala Asn Lys Ser Ser Gln Leu Val Thr Phe His Ile Asp Leu Asn 145 150 155 160 Ser Val Asp Ile Leu Asp Lys Leu Thr Thr Thr Ser His Leu Ala Thr 165 170 175 Pro Gly Asn Val Ser Ile Glu Cys Leu Pro Gly Ser Ser Pro Cys Thr 180 185 190 Asp Ala Leu Thr Cys Ile Lys Ala Asp Leu Phe Cys Asp Gly Glu Val 195 200 205 Asn Cys Pro Asp Gly Ser Asp Glu Asp Asn Lys Met Cys Ala Thr Val 210 215 220 Cys Asp Gly Arg Phe Leu Leu Thr Gly Ser Ser Gly Ser Phe Gln Ala 225 230 235 240 Thr His Tyr Pro Lys Pro Ser Glu Thr Ser Val Val Cys Gln Trp Ile 245 250 255 Ile Arg Val Asn Gln Gly Leu Ser Ile Lys Leu Ser Phe Asp Asp Phe 260 265 270 Asn Thr Tyr Tyr Thr Asp Ile Leu Asp Ile Tyr Glu Gly Val Gly Ser 275 280 285 Ser Lys Ile Leu Arg Ala Ser Ile Trp Glu Thr Asn Pro Gly Thr Ile 290 295 300 Arg Ile Phe Ser Asn Gln Val Thr Ala Thr Phe Leu Ile Glu Ser Asp 305 310 315 320 Glu Ser Asp Tyr Val Gly Phe Asn Ala Thr Tyr Thr Ala Phe Asn Ser 325 330 335 Ser Glu Leu Asn Asn Tyr Glu Lys Ile Asn Cys Asn Phe Glu Asp Gly 340 345 350 Phe Cys Phe Trp Val Gln Asp Leu Asn Asp Asp Asn Glu Trp Glu Arg 355 360 365 Ile Gln Gly Ser Thr Phe Ser Pro Phe Thr Gly Pro Asn Phe Asp His 370 375 380 Thr Phe Gly Asn Ala Ser Gly Phe Tyr Ile Ser Thr Pro Thr Gly Pro 385 390 395 400 Gly Gly Arg Gln Glu Arg Val Gly Leu Leu Ser Leu Pro Leu Asp Pro 405 410 415 Thr Leu Glu Pro Ala Cys Leu Ser Phe Trp Tyr His Met Tyr Gly Glu 420 425 430 Asn Val His Lys Leu Ser Ile Asn Ile Ser Asn Asp Gln Asn Met Glu 435 440 445 Lys Thr Val Phe Gln Lys Glu Gly Asn Tyr Gly Asp Asn Trp Asn Tyr 450 455 460 Gly Gln Val Thr Leu Asn Glu Thr Val Lys Phe Lys Val Ala Phe Asn 465 470 475 480 Ala Phe Lys Asn Lys Ile Leu Ser Asp Ile Ala Leu Asp Asp Ile Ser 485 490 495 Leu Thr Tyr Gly Ile Cys Asn Gly Ser Leu Tyr Pro Glu Pro Thr Leu 500 505 510 Val Pro Thr Pro Pro Pro Glu Leu Pro Thr Asp Cys Gly Gly Pro Phe 515 520 525 Glu Leu Trp Glu Pro Asn Thr Thr Phe Ser Ser Thr Asn Phe Pro Asn 530 535 540 Ser Tyr Pro Asn Leu Ala Phe Cys Val Trp Ile Leu Asn Ala Gln Lys 545 550 555 560 Gly Lys Asn Ile Gln Leu His Phe Gln Glu Phe Asp Leu Glu Asn Ile 565 570 575 Asn Asp Val Val Glu Ile Arg Asp Gly Glu Glu Ala Asp Ser Leu Leu 580 585 590 Leu Ala Val Tyr Thr Gly Pro Gly Pro Val Lys Asp Val Phe Ser Thr 595 600 605 Thr Asn Arg Met Thr Val Leu Leu Ile Thr Asn Asp Val Leu Ala Arg 610 615 620 Gly Gly Phe Lys Ala Asn Phe Thr Thr Gly Tyr His Leu Gly Ile Pro 625 630 635 640 Glu Pro Cys Lys Ala Asp His Phe Gln Cys Lys Asn Gly Glu Cys Val 645 650 655 Pro Leu Val Asn Leu Cys Asp Gly His Leu His Cys Glu Asp Gly Ser 660 665 670 Asp Glu Ala Asp Cys Val Arg Phe Phe Asn Gly Thr Thr Asn Asn Asn 675 680 685 Gly Leu Val Arg Phe Arg Ile Gln Ser Ile Trp His Thr Ala Cys Ala 690 695 700 Glu Asn Trp Thr Thr Gln Ile Ser Asn Asp Val Cys Gln Leu Leu Gly 705 710 715 720 Leu Gly Ser Gly Asn Ser Ser Lys Pro Ile Phe Ser Thr Asp Gly Gly 725 730 735 Pro Phe Val Lys Leu Asn Thr Ala Pro Asp Gly His Leu Ile Leu Thr 740 745 750 Pro Ser Gln Gln Cys Leu Gln Asp Ser Leu Ile Arg Leu Gln Cys Asn 755 760 765 His Lys Ser Cys Gly Lys Lys Leu Ala Ala Gln Asp Ile Thr Pro Lys 770 775 780 Ile Val Gly Gly Ser Asn Ala Lys Glu Gly Ala Trp Pro Trp Val Val 785 790 795 800 Gly Leu Tyr Tyr Gly Gly Arg Leu Leu Cys Gly Ala Ser Leu Val Ser 805 810 815 Ser Asp Trp Leu Val Ser Ala Ala His Cys Val Tyr Gly Arg Asn Leu 820 825 830 Glu Pro Ser Lys Trp Thr Ala Ile Leu Gly Leu His Met Lys Ser Asn 835 840 845 Leu Thr Ser Pro Gln Thr Val Pro Arg Leu Ile Asp Glu Ile Val Ile 850 855 860 Asn Pro His Tyr Asn Arg Arg Arg Lys Asp Asn Asp Ile Ala Met Met 865 870 875 880 His Leu Glu Phe Lys Val Asn Tyr Thr Asp Tyr Ile Gln Pro Ile Cys 885 890 895 Leu Pro Glu Glu Asn Gln Val Phe Pro Pro Gly Arg Asn Cys Ser Ile 900 905 910 Ala Gly Trp Gly Thr Val Val Tyr Gln Gly Thr Thr Ala Asn Ile Leu 915 920 925 Gln Glu Ala Asp Val Pro Leu Leu Ser Asn Glu Arg Cys Gln Gln Gln 930 935 940 Met Pro Glu Tyr Asn Ile Thr Glu Asn Met Ile Cys Ala Gly Tyr Glu 945 950 955 960 Glu Gly Gly Ile Asp Ser Cys Gln Gly Asp Ser Gly Gly Pro Leu Met 965 970 975 Cys Gln Glu Asn Asn Arg Trp Phe Leu Ala Gly Val Thr Ser Phe Gly 980 985 990 Tyr Lys Cys Ala Leu Pro Asn Arg Pro Gly Val Tyr Ala Arg Val Ser 995 1000 1005 Arg Phe Thr Glu Trp Ile Gln Ser Phe Leu His 1010 1015 <210> SEQ ID NO 32 <211> LENGTH: 1500 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (62)...(1318) <223> OTHER INFORMATION: Nucleotide sequence encoding human airway trypsin-like protease <300> PUBLICATION INFORMATION: <308> DATABASE ACCESSION NUMBER: GenBank AB002134 <309> DATABASE ENTRY DATE: 1998-06-04 <400> SEQUENCE: 32 gagtgggaat ctcaaagcag ttgagtaggc agaaaaaaga acctcttcat taaggattaa 60 a atg tat agg cca gca cgt gta act tcg act tca aga ttt ctg aat cca 109 Met Tyr Arg Pro Ala Arg Val Thr Ser Thr Ser Arg Phe Leu Asn Pro 1 5 10 15 tat gta gta tgt ttc att gtc gtc gca ggg gta gtg atc ctg gca gtc 157 Tyr Val Val Cys Phe Ile Val Val Ala Gly Val Val Ile Leu Ala Val 20 25 30 acc ata gct cta ctt gtt tac ttt tta gct ttt gat caa aaa tct tac 205 Thr Ile Ala Leu Leu Val Tyr Phe Leu Ala Phe Asp Gln Lys Ser Tyr 35 40 45 ttt tat agg agc agt ttt caa ctc cta aat gtt gaa tat aat agt cag 253 Phe Tyr Arg Ser Ser Phe Gln Leu Leu Asn Val Glu Tyr Asn Ser Gln 50 55 60 tta aat tca cca gct aca cag gaa tac agg act ttg agt gga aga att 301 Leu Asn Ser Pro Ala Thr Gln Glu Tyr Arg Thr Leu Ser Gly Arg Ile 65 70 75 80 gaa tct ctg att act aaa aca ttc aaa gaa tca aat tta aga aat cag 349Glu Ser Leu Ile Thr Lys Thr Phe Lys Glu Ser Asn Leu Arg Asn Gln 85 90 95 ttc atc aga gct cat gtt gcc aaa ctg agg caa gat ggt agt ggt gtg 397 Phe Ile Arg Ala His Val Ala Lys Leu Arg Gln Asp Gly Ser Gly Val 100 105 110 aga gcg gat gtt gtc atg aaa ttt caa ttc act aga aat aac aat gga 445Arg Ala Asp Val Val Met Lys Phe Gln Phe Thr Arg Asn Asn Asn Gly 115 120 125 gca tca atg aaa agc aga att gag tct gtt tta cga caa atg ctg aat 493 Ala Ser Met Lys Ser Arg Ile Glu Ser Val Leu Arg Gln Met Leu Asn 130 135 140 aac tct gga aac ctg gaa ata aac cct tca act gag ata aca tca ctt 541Asn Ser Gly Asn Leu Glu Ile Asn Pro Ser Thr Glu Ile Thr Ser Leu 145 150 155 160 act gac cag gct gca gca aat tgg ctt att aat gaa tgt ggg gcc ggt 589 Thr Asp Gln Ala Ala Ala Asn Trp Leu Ile Asn Glu Cys Gly Ala Gly 165 170 175 cca gac cta ata aca ttg tct gag cag aga atc ctt gga ggc act gag 637 Pro Asp Leu Ile Thr Leu Ser Glu Gln Arg Ile Leu Gly Gly Thr Glu 180 185 190 gct gag gag gga agc tgg ccg tgg caa gtc agt ctg cgg ctc aat aat 685 Ala Glu Glu Gly Ser Trp Pro Trp Gln Val Ser Leu Arg Leu Asn Asn 195 200 205 gcc cac cac tgt gga ggc agc ctg atc aat aac atg tgg atc ctg aca 733 Ala His His Cys Gly Gly Ser Leu Ile Asn Asn Met Trp Ile Leu Thr 210 215 220 gca gct cac tgc ttc aga agc aac tct aat cct cgt gac tgg att gcc 781 Ala Ala His Cys Phe Arg Ser Asn Ser Asn Pro Arg Asp Trp Ile Ala 225 230 235 240 acg tct ggt att tcc aca aca ttt cct aaa cta aga atg aga gta aga 829 Thr Ser Gly Ile Ser Thr Thr Phe Pro Lys Leu Arg Met Arg Val Arg 245 250 255 aat att tta att cat aac aat tat aaa tct gca act cat gaa aat gac 877 Asn Ile Leu Ile His Asn Asn Tyr Lys Ser Ala Thr His Glu Asn Asp 260 265 270 att gca ctt gtg aga ctt gag aac agt gtc acc ttt acc aaa gat atc 925 Ile Ala Leu Val Arg Leu Glu Asn Ser Val Thr Phe Thr Lys Asp Ile 275 280 285 cat agt gtg tgt ctc cca gct gct acc cag aat att cca cct ggc tct 973 His Ser Val Cys Leu Pro Ala Ala Thr Gln Asn Ile Pro Pro Gly Ser 290 295 300 act gct tat gta aca gga tgg ggc gct caa gaa tat gct ggc cac aca 1021 Thr Ala Tyr Val Thr Gly Trp Gly Ala Gln Glu Tyr Ala Gly His Thr 305 310 315 320 gtt cca gag cta agg caa gga cag gtc aga ata ata agt aat gat gta 1069 Val Pro Glu Leu Arg Gln Gly Gln Val Arg Ile Ile Ser Asn Asp Val 325 330 335 tgt aat gca cca cat agt tat aat gga gcc atc ttg tct gga atg ctg 1117 Cys Asn Ala Pro His Ser Tyr Asn Gly Ala Ile Leu Ser Gly Met Leu 340 345 350 tgt gct gga gta cct caa ggt gga gtg gac gca tgt cag ggt gac tct 1165 Cys Ala Gly Val Pro Gln Gly Gly Val Asp Ala Cys Gln Gly Asp Ser 355 360 365 ggt ggc cca cta gta caa gaa gac tca cgg cgg ctt tgg ttt att gtg 1213 Gly Gly Pro Leu Val Gln Glu Asp Ser Arg Arg Leu Trp Phe Ile Val 370 375 380 ggg ata gta agc tgg gga gat cag tgt ggc ctg ccg gat aag cca gga 1261 Gly Ile Val Ser Trp Gly Asp Gln Cys Gly Leu Pro Asp Lys Pro Gly 385 390 395 400 gtg tat act cga gtg aca gcc tac ctt gac tgg att agg caa caa act 1309 Val Tyr Thr Arg Val Thr Ala Tyr Leu Asp Trp Ile Arg Gln Gln Thr 405 410 415 ggg atc tag tgcaacaagt gcatccctgt tgcaaagtct gtatgcaggt 1358 Gly Ile * gtgcctgtct taaattccaa agctttacat ttcaactgaa aaagaaacta gaaatgtcct 1418 aatttaacat cttgttacat aaatatggtt taacaaacac tgtttaacct ttctttatta 1478 ttaaaggttt tctattttct cc 1500 <210> SEQ ID NO 33 <211> LENGTH: 418 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 33 Met Tyr Arg Pro Ala Arg Val Thr Ser Thr Ser Arg Phe Leu Asn Pro 1 5 10 15 Tyr Val Val Cys Phe Ile Val Val Ala Gly Val Val Ile Leu Ala Val 20 25 30 Thr Ile Ala Leu Leu Val Tyr Phe Leu Ala Phe Asp Gln Lys Ser Tyr 35 40 45 Phe Tyr Arg Ser Ser Phe Gln Leu Leu Asn Val Glu Tyr Asn Ser Gln 50 55 60 Leu Asn Ser Pro Ala Thr Gln Glu Tyr Arg Thr Leu Ser Gly Arg Ile 65 70 75 80 Glu Ser Leu Ile Thr Lys Thr Phe Lys Glu Ser Asn Leu Arg Asn Gln 85 90 95 Phe Ile Arg Ala His Val Ala Lys Leu Arg Gln Asp Gly Ser Gly Val 100 105 110 Arg Ala Asp Val Val Met Lys Phe Gln Phe Thr Arg Asn Asn Asn Gly 115 120 125 Ala Ser Met Lys Ser Arg Ile Glu Ser Val Leu Arg Gln Met Leu Asn 130 135 140 Asn Ser Gly Asn Leu Glu Ile Asn Pro Ser Thr Glu Ile Thr Ser Leu 145 150 155 160 Thr Asp Gln Ala Ala Ala Asn Trp Leu Ile Asn Glu Cys Gly Ala Gly 165 170 175 Pro Asp Leu Ile Thr Leu Ser Glu Gln Arg Ile Leu Gly Gly Thr Glu 180 185 190 Ala Glu Glu Gly Ser Trp Pro Trp Gln Val Ser Leu Arg Leu Asn Asn 195 200 205 Ala His His Cys Gly Gly Ser Leu Ile Asn Asn Met Trp Ile Leu Thr 210 215 220 Ala Ala His Cys Phe Arg Ser Asn Ser Asn Pro Arg Asp Trp Ile Ala 225 230 235 240 Thr Ser Gly Ile Ser Thr Thr Phe Pro Lys Leu Arg Met Arg Val Arg 245 250 255 Asn Ile Leu Ile His Asn Asn Tyr Lys Ser Ala Thr His Glu Asn Asp 260 265 270 Ile Ala Leu Val Arg Leu Glu Asn Ser Val Thr Phe Thr Lys Asp Ile 275 280 285 His Ser Val Cys Leu Pro Ala Ala Thr Gln Asn Ile Pro Pro Gly Ser 290 295 300 Thr Ala Tyr Val Thr Gly Trp Gly Ala Gln Glu Tyr Ala Gly His Thr 305 310 315 320 Val Pro Glu Leu Arg Gln Gly Gln Val Arg Ile Ile Ser Asn Asp Val 325 330 335 Cys Asn Ala Pro His Ser Tyr Asn Gly Ala Ile Leu Ser Gly Met Leu 340 345 350 Cys Ala Gly Val Pro Gln Gly Gly Val Asp Ala Cys Gln Gly Asp Ser 355 360 365 Gly Gly Pro Leu Val Gln Glu Asp Ser Arg Arg Leu Trp Phe Ile Val 370 375 380 Gly Ile Val Ser Trp Gly Asp Gln Cys Gly Leu Pro Asp Lys Pro Gly 385 390 395 400 Val Tyr Thr Arg Val Thr Ala Tyr Leu Asp Trp Ile Arg Gln Gln Thr 405 410 415 Gly Ile <210> SEQ ID NO 34 <211> LENGTH: 1783 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (246)...(1499) <223> OTHER INFORMATION: Nucleic acid encoding human hepsin <300> PUBLICATION INFORMATION: <308> DATABASE ACCESSION NUMBER: GenBank M18930 <309> DATABASE ENTRY DATE: 1993-06-11 <400> SEQUENCE: 34 tcgagcccgc tttccaggga ccctacctga gggcccacag gtgaggcagc ctggcctagc 60 aggccccacg ccaccgcctc tgcctccagg ccgcccgctg ctgcggggcc accatgctcc 120 tgcccaggcc tggagactga cccgaccccg gcactacctc gaggctccgc ccccacctgc 180 tggaccccag ggtcccaccc tggcccagga ggtcagccag ggaatcatta acaagaggca 240 gtgac atg gcg cag aag gag ggt ggc cgg act gtg cca tgc tgc tcc aga 290 Met Ala Gln Lys Glu Gly Gly Arg Thr Val Pro Cys Cys Ser Arg 1 5 10 15 ccc aag gtg gca gct ctc act gcg ggg acc ctg cta ctt ctg aca gcc 338Pro Lys Val Ala Ala Leu Thr Ala Gly Thr Leu Leu Leu Leu Thr Ala 20 25 30 atc ggg gcg gca tcc tgg gcc att gtg gct gtt ctc ctc agg agt gac 386Ile Gly Ala Ala Ser Trp Ala Ile Val Ala Val Leu Leu Arg Ser Asp 35 40 45 cag gag ccg ctg tac cca gtg cag gtc agc tct gcg gac gct cgg ctc 434Gln Glu Pro Leu Tyr Pro Val Gln Val Ser Ser Ala Asp Ala Arg Leu 50 55 60 atg gtc ttt gac aag acg gaa ggg acg tgg cgg ctg ctg tgc tcc tcg 482Met Val Phe Asp Lys Thr Glu Gly Thr Trp Arg Leu Leu Cys Ser Ser 65 70 75 cgc tcc aac gcc agg gta gcc gga ctc agc tgc gag gag atg ggc ttc 530Arg Ser Asn Ala Arg Val Ala Gly Leu Ser Cys Glu Glu Met Gly Phe 80 85 90 95 ctc agg gca ctg acc cac tcc gag ctg gac gtg cga acg gcg ggc gcc 578 Leu Arg Ala Leu Thr His Ser Glu Leu Asp Val Arg Thr Ala Gly Ala 100 105 110 aat ggc acg tcg ggc ttc ttc tgt gtg gac gag ggg agg ctg ccc cac 626 Asn Gly Thr Ser Gly Phe Phe Cys Val Asp Glu Gly Arg Leu Pro His 115 120 125 acc cag agg ctg ctg gag gtc atc tcc gtg tgt gat tgc ccc aga ggc 674 Thr Gln Arg Leu Leu Glu Val Ile Ser Val Cys Asp Cys Pro Arg Gly 130 135 140 cgt ttc ttg gcc gcc atc tgc caa gac tgt ggc cgc agg aag ctg ccc 722 Arg Phe Leu Ala Ala Ile Cys Gln Asp Cys Gly Arg Arg Lys Leu Pro 145 150 155 gtg gac cgc atc gtg gga ggc cgg gac acc agc ttg ggc cgg tgg ccg 770 Val Asp Arg Ile Val Gly Gly Arg Asp Thr Ser Leu Gly Arg Trp Pro 160 165 170 175 tgg caa gtc agc ctt cgc tat gat gga gca cac ctc tgt ggg gga tcc 818 Trp Gln Val Ser Leu Arg Tyr Asp Gly Ala His Leu Cys Gly Gly Ser 180 185 190 ctg ctc tcc ggg gac tgg gtg ctg aca gcc gcc cac tgc ttc ccg gag 866 Leu Leu Ser Gly Asp Trp Val Leu Thr Ala Ala His Cys Phe Pro Glu 195 200 205 cgg aac cgg gtc ctg tcc cga tgg cga gtg ttt gcc ggt gcc gtg gcc 914 Arg Asn Arg Val Leu Ser Arg Trp Arg Val Phe Ala Gly Ala Val Ala 210 215 220 cag gcc tct ccc cac ggt ctg cag ctg ggg gtg cag gct gtg gtc tac 962 Gln Ala Ser Pro His Gly Leu Gln Leu Gly Val Gln Ala Val Val Tyr 225 230 235 cac ggg ggc tat ctt ccc ttt cgg gac ccc aac agc gag gag aac agc 1010 His Gly Gly Tyr Leu Pro Phe Arg Asp Pro Asn Ser Glu Glu Asn Ser 240 245 250 255 aac gat att gcc ctg gtc cac ctc tcc agt ccc ctg ccc ctc aca gaa 1058 Asn Asp Ile Ala Leu Val His Leu Ser Ser Pro Leu Pro Leu Thr Glu 260 265 270 tac atc cag cct gtg tgc ctc cca gct gcc ggc cag gcc ctg gtg gat 1106 Tyr Ile Gln Pro Val Cys Leu Pro Ala Ala Gly Gln Ala Leu Val Asp 275 280 285 ggc aag atc tgt acc gtg acg ggc tgg ggc aac acg cag tac tat ggc 1154 Gly Lys Ile Cys Thr Val Thr Gly Trp Gly Asn Thr Gln Tyr Tyr Gly 290 295 300 caa cag gcc ggg gta ctc cag gag gct cga gtc ccc ata atc agc aat 1202 Gln Gln Ala Gly Val Leu Gln Glu Ala Arg Val Pro Ile Ile Ser Asn 305 310 315 gat gtc tgc aat ggc gct gac ttc tat gga aac cag atc aag ccc aag 1250 Asp Val Cys Asn Gly Ala Asp Phe Tyr Gly Asn Gln Ile Lys Pro Lys 320 325 330 335 atg ttc tgt gct ggc tac ccc gag ggt ggc att gat gcc tgc cag ggc 1298 Met Phe Cys Ala Gly Tyr Pro Glu Gly Gly Ile Asp Ala Cys Gln Gly 340 345 350 gac agc ggt ggt ccc ttt gtg tgt gag gac agc atc tct cgg acg cca 1346 Asp Ser Gly Gly Pro Phe Val Cys Glu Asp Ser Ile Ser Arg Thr Pro 355 360 365 cgt tgg cgg ctg tgt ggc att gtg agt tgg ggc act ggc tgt gcc ctg 1394 Arg Trp Arg Leu Cys Gly Ile Val Ser Trp Gly Thr Gly Cys Ala Leu 370 375 380 gcc cag aag cca ggc gtc tac acc aaa gtc agt gac ttc cgg gag tgg 1442 Ala Gln Lys Pro Gly Val Tyr Thr Lys Val Ser Asp Phe Arg Glu Trp 385 390 395 atc ttc cag gcc ata aag act cac tcc gaa gcc agc ggc atg gtg acc 1490 Ile Phe Gln Ala Ile Lys Thr His Ser Glu Ala Ser Gly Met Val Thr 400 405 410 415 cag ctc tga ccggtggctt ctcgctgcgc agcctccagg gcccgaggtg 1539 Gln Leu * atcccggtgg tgggatccac gctgggccga ggatgggacg tttttcttct tgggcccggt 1599 ccacaggtcc aaggacaccc tccctccagg gtcctctctt ccacagtggc gggcccactc 1659 agccccgaga ccacccaacc tcaccctcct gacccccatg taaatattgt tctgctgtct 1719 gggactcctg tctaggtgcc cctgatgatg ggatgctctt taaataataa agatggtttt 1779 gatt 1783 <210> SEQ ID NO 35 <211> LENGTH: 417 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 35 Met Ala Gln Lys Glu Gly Gly Arg Thr Val Pro Cys Cys Ser Arg Pro 1 5 10 15 Lys Val Ala Ala Leu Thr Ala Gly Thr Leu Leu Leu Leu Thr Ala Ile 20 25 30 Gly Ala Ala Ser Trp Ala Ile Val Ala Val Leu Leu Arg Ser Asp Gln 35 40 45 Glu Pro Leu Tyr Pro Val Gln Val Ser Ser Ala Asp Ala Arg Leu Met 50 55 60 Val Phe Asp Lys Thr Glu Gly Thr Trp Arg Leu Leu Cys Ser Ser Arg 65 70 75 80 Ser Asn Ala Arg Val Ala Gly Leu Ser Cys Glu Glu Met Gly Phe Leu 85 90 95 Arg Ala Leu Thr His Ser Glu Leu Asp Val Arg Thr Ala Gly Ala Asn 100 105 110 Gly Thr Ser Gly Phe Phe Cys Val Asp Glu Gly Arg Leu Pro His Thr 115 120 125 Gln Arg Leu Leu Glu Val Ile Ser Val Cys Asp Cys Pro Arg Gly Arg 130 135 140 Phe Leu Ala Ala Ile Cys Gln Asp Cys Gly Arg Arg Lys Leu Pro Val 145 150 155 160 Asp Arg Ile Val Gly Gly Arg Asp Thr Ser Leu Gly Arg Trp Pro Trp 165 170 175 Gln Val Ser Leu Arg Tyr Asp Gly Ala His Leu Cys Gly Gly Ser Leu 180 185 190 Leu Ser Gly Asp Trp Val Leu Thr Ala Ala His Cys Phe Pro Glu Arg 195 200 205 Asn Arg Val Leu Ser Arg Trp Arg Val Phe Ala Gly Ala Val Ala Gln 210 215 220 Ala Ser Pro His Gly Leu Gln Leu Gly Val Gln Ala Val Val Tyr His 225 230 235 240 Gly Gly Tyr Leu Pro Phe Arg Asp Pro Asn Ser Glu Glu Asn Ser Asn 245 250 255 Asp Ile Ala Leu Val His Leu Ser Ser Pro Leu Pro Leu Thr Glu Tyr 260 265 270 Ile Gln Pro Val Cys Leu Pro Ala Ala Gly Gln Ala Leu Val Asp Gly 275 280 285 Lys Ile Cys Thr Val Thr Gly Trp Gly Asn Thr Gln Tyr Tyr Gly Gln 290 295 300 Gln Ala Gly Val Leu Gln Glu Ala Arg Val Pro Ile Ile Ser Asn Asp 305 310 315 320 Val Cys Asn Gly Ala Asp Phe Tyr Gly Asn Gln Ile Lys Pro Lys Met 325 330 335 Phe Cys Ala Gly Tyr Pro Glu Gly Gly Ile Asp Ala Cys Gln Gly Asp 340 345 350 Ser Gly Gly Pro Phe Val Cys Glu Asp Ser Ile Ser Arg Thr Pro Arg 355 360 365 Trp Arg Leu Cys Gly Ile Val Ser Trp Gly Thr Gly Cys Ala Leu Ala 370 375 380 Gln Lys Pro Gly Val Tyr Thr Lys Val Ser Asp Phe Arg Glu Trp Ile 385 390 395 400 Phe Gln Ala Ile Lys Thr His Ser Glu Ala Ser Gly Met Val Thr Gln 405 410 415 Leu <210> SEQ ID NO 36 <211> LENGTH: 2479 <212> TYPE: DNA <213> ORGANISM: Homo sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (57)...(1535) <223> OTHER INFORMATION: Nucleotide sequence encoding human serine protease (TMPRS2) <300> PUBLICATION INFORMATION: <308> DATABASE ACCESSION NUMBER: GenBank U75329 <309> DATABASE ENTRY DATE: 1997-10-10 <400> SEQUENCE: 36 gtcatattga acattccaga tacctatcat tactcgatgc tgttgataac agcaag atg 59 Met 1 gct ttg aac tca ggg tca cca cca gct att gga cct tac tat gaa aac 107 Ala Leu Asn Ser Gly Ser Pro Pro Ala Ile Gly Pro Tyr Tyr Glu Asn 5 10 15 cat gga tac caa ccg gaa aac ccc tat ccc gca cag ccc act gtg gtc 155 His Gly Tyr Gln Pro Glu Asn Pro Tyr Pro Ala Gln Pro Thr Val Val 20 25 30 ccc act gtc tac gag gtg cat ccg gct cag tac tac ccg tcc ccc gtg 203 Pro Thr Val Tyr Glu Val His Pro Ala Gln Tyr Tyr Pro Ser Pro Val 35 40 45 ccc cag tac gcc ccg agg gtc ctg acg cag gct tcc aac ccc gtc gtc 251 Pro Gln Tyr Ala Pro Arg Val Leu Thr Gln Ala Ser Asn Pro Val Val 50 55 60 65 tgc acg cag ccc aaa tcc cca tcc ggg aca gtg tgc acc tca aag act 299Cys Thr Gln Pro Lys Ser Pro Ser Gly Thr Val Cys Thr Ser Lys Thr 70 75 80 aag aaa gca ctg tgc atc acc ttg acc ctg ggg acc ttc ctc gtg gga 347Lys Lys Ala Leu Cys Ile Thr Leu Thr Leu Gly Thr Phe Leu Val Gly 85 90 95 gct gcg ctg gcc gct ggc cta ctc tgg aag ttc atg ggc agc aag tgc 395Ala Ala Leu Ala Ala Gly Leu Leu Trp Lys Phe Met Gly Ser Lys Cys 100 105 110 tcc aac tct ggg ata gag tgc gac tcc tca ggt acc tgc atc aac ccc 443 Ser Asn Ser Gly Ile Glu Cys Asp Ser Ser Gly Thr Cys Ile Asn Pro 115 120 125 tct aac tgg tgt gat ggc gtg tca cac tgc ccc ggc ggg gag gac gag 491 Ser Asn Trp Cys Asp Gly Val Ser His Cys Pro Gly Gly Glu Asp Glu 130 135 140 145 aat cgg tgt gtt cgc ctc tac gga cca aac ttc atc ctt cag atg tac 539Asn Arg Cys Val Arg Leu Tyr Gly Pro Asn Phe Ile Leu Gln Met Tyr 150 155 160 tca tct cag agg aag tcc tgg cac cct gtg tgc caa gac gac tgg aac 587 Ser Ser Gln Arg Lys Ser Trp His Pro Val Cys Gln Asp Asp Trp Asn 165 170 175 gag aac tac ggg cgg gcg gcc tgc agg gac atg ggc tat aag aat aat 635 Glu Asn Tyr Gly Arg Ala Ala Cys Arg Asp Met Gly Tyr Lys Asn Asn 180 185 190 ttt tac tct agc caa gga ata gtg gat gac agc gga tcc acc agc ttt 683 Phe Tyr Ser Ser Gln Gly Ile Val Asp Asp Ser Gly Ser Thr Ser Phe 195 200 205 atg aaa ctg aac aca agt gcc ggc aat gtc gat atc tat aaa aaa ctg 731 Met Lys Leu Asn Thr Ser Ala Gly Asn Val Asp Ile Tyr Lys Lys Leu 210 215 220 225 tac cac agt gat gcc tgt tct tca aaa gca gtg gtt tct tta cgc tgt 779 Tyr His Ser Asp Ala Cys Ser Ser Lys Ala Val Val Ser Leu Arg Cys 230 235 240 tta gcc tgc ggg gtc aac ttg aac tca agc cgc cag agc agg atc gtg 827 Leu Ala Cys Gly Val Asn Leu Asn Ser Ser Arg Gln Ser Arg Ile Val 245 250 255 ggc ggt gag agc gcg ctc ccg ggg gcc tgg ccc tgg cag gtc agc ctg 875 Gly Gly Glu Ser Ala Leu Pro Gly Ala Trp Pro Trp Gln Val Ser Leu 260 265 270 cac gtc cag aac gtc cac gtg tgc gga ggc tcc atc atc acc ccc gag 923 His Val Gln Asn Val His Val Cys Gly Gly Ser Ile Ile Thr Pro Glu 275 280 285 tgg atc gtg aca gcc gcc cac tgc gtg gaa aaa cct ctt aac aat cca 971 Trp Ile Val Thr Ala Ala His Cys Val Glu Lys Pro Leu Asn Asn Pro 290 295 300 305 tgg cat tgg acg gca ttt gcg ggg att ttg aga caa tct ttc atg ttc 1019 Trp His Trp Thr Ala Phe Ala Gly Ile Leu Arg Gln Ser Phe Met Phe 310 315 320 tat gga gcc gga tac caa gta caa aaa gtg att tct cat cca aat tat 1067 Tyr Gly Ala Gly Tyr Gln Val Gln Lys Val Ile Ser His Pro Asn Tyr 325 330 335 gac tcc aag acc aag aac aat gac att gcg ctg atg aag ctg cag aag 1115 Asp Ser Lys Thr Lys Asn Asn Asp Ile Ala Leu Met Lys Leu Gln Lys 340 345 350 cct ctg act ttc aac gac cta gtg aaa cca gtg tgt ctg ccc aac cca 1163 Pro Leu Thr Phe Asn Asp Leu Val Lys Pro Val Cys Leu Pro Asn Pro 355 360 365 ggc atg atg ctg cag cca gaa cag ctc tgc tgg att tcc ggg tgg ggg 1211 Gly Met Met Leu Gln Pro Glu Gln Leu Cys Trp Ile Ser Gly Trp Gly 370 375 380 385 gcc acc gag gag aaa ggg aag acc tca gaa gtg ctg aac gct gcc aag 1259 Ala Thr Glu Glu Lys Gly Lys Thr Ser Glu Val Leu Asn Ala Ala Lys 390 395 400 gtg ctt ctc att gag aca cag aga tgc aac agc aga tat gtc tat gac 1307 Val Leu Leu Ile Glu Thr Gln Arg Cys Asn Ser Arg Tyr Val Tyr Asp 405 410 415 aac ctg atc aca cca gcc atg atc tgt gcc ggc ttc ctg cag ggg aac 1355 Asn Leu Ile Thr Pro Ala Met Ile Cys Ala Gly Phe Leu Gln Gly Asn 420 425 430 gtc gat tct tgc cag ggt gac agt gga ggg cct ctg gtc act tcg aac 1403 Val Asp Ser Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Thr Ser Asn 435 440 445 aac aat atc tgg tgg ctg ata ggg gat aca agc tgg ggt tct ggc tgt 1451 Asn Asn Ile Trp Trp Leu Ile Gly Asp Thr Ser Trp Gly Ser Gly Cys 450 455 460 465 gcc aaa gct tac aga cca gga gtg tac ggg aat gtg atg gta ttc acg 1499 Ala Lys Ala Tyr Arg Pro Gly Val Tyr Gly Asn Val Met Val Phe Thr 470 475 480 gac tgg att tat cga caa atg aag gca aac ggc taa tccacatggt 1545 Asp Trp Ile Tyr Arg Gln Met Lys Ala Asn Gly * 485 490 cttcgtcctt gacgtcgttt tacaagaaaa caatggggct ggttttgctt ccccgtgcat 1605 gatttactct tagagatgat tcagaggtca cttcattttt attaaacagt gaacttgtct 1665 ggctttggca ctctctgcca tactgtgcag gctgcagtgg ctcccctgcc cagcctgctc 1725 tccctaaccc cttgtccgca aggggtgatg gccggctggt tgtgggcact ggcggtcaat 1785 tgtggaagga agagggttgg aggctgcccc cattgagatc ttcctgctga gtcctttcca 1845 ggggccaatt ttggatgagc atggagctgt cacttctcag ctgctggatg acttgagatg 1905 aaaaaggaga gacatggaaa gggagacagc caggtggcac ctgcagcggc tgccctctgg 1965 ggccacttgg tagtgtcccc agcctacttc acaaggggat tttgctgatg ggttcttaga 2025 gccttagcag ccctggatgg tggccagaaa taaagggacc agcccttcat gggtggtgac 2085 gtggtagtca cttgtaaggg gaacagaaac atttttgttc ttatggggtg agaatataga 2145 cagtgccctt ggtgcgaggg aagcaattga aaaggaactt gccctgagca ctcctggtgc 2205 aggtctccac ctgcacattg ggtggggctc ctgggaggga gactcagcct tcctcctcat 2265 cctccctgac cctgctccta gcaccctgga gagtgaatgc cccttggtcc ctggcagggc 2325 gccaagtttg gcaccatgtc ggcctcttca ggcctgatag tcattggaaa ttgaggtcca 2385 tgggggaaat caaggatgct cagtttaagg tacactgttt ccatgttatg tttctacaca 2445 ttgatggtgg tgaccctgag ttcaaagcca tctt 2479 <210> SEQ ID NO 37 <211> LENGTH: 492 <212> TYPE: PRT <213> ORGANISM: Homo sapien <400> SEQUENCE: 37 Met Ala Leu Asn Ser Gly Ser Pro Pro Ala Ile Gly Pro Tyr Tyr Glu 1 5 10 15 Asn His Gly Tyr Gln Pro Glu Asn Pro Tyr Pro Ala Gln Pro Thr Val 20 25 30 Val Pro Thr Val Tyr Glu Val His Pro Ala Gln Tyr Tyr Pro Ser Pro 35 40 45 Val Pro Gln Tyr Ala Pro Arg Val Leu Thr Gln Ala Ser Asn Pro Val 50 55 60 Val Cys Thr Gln Pro Lys Ser Pro Ser Gly Thr Val Cys Thr Ser Lys 65 70 75 80 Thr Lys Lys Ala Leu Cys Ile Thr Leu Thr Leu Gly Thr Phe Leu Val 85 90 95 Gly Ala Ala Leu Ala Ala Gly Leu Leu Trp Lys Phe Met Gly Ser Lys 100 105 110 Cys Ser Asn Ser Gly Ile Glu Cys Asp Ser Ser Gly Thr Cys Ile Asn 115 120 125 Pro Ser Asn Trp Cys Asp Gly Val Ser His Cys Pro Gly Gly Glu Asp 130 135 140 Glu Asn Arg Cys Val Arg Leu Tyr Gly Pro Asn Phe Ile Leu Gln Met 145 150 155 160 Tyr Ser Ser Gln Arg Lys Ser Trp His Pro Val Cys Gln Asp Asp Trp 165 170 175 Asn Glu Asn Tyr Gly Arg Ala Ala Cys Arg Asp Met Gly Tyr Lys Asn 180 185 190 Asn Phe Tyr Ser Ser Gln Gly Ile Val Asp Asp Ser Gly Ser Thr Ser 195 200 205 Phe Met Lys Leu Asn Thr Ser Ala Gly Asn Val Asp Ile Tyr Lys Lys 210 215 220 Leu Tyr His Ser Asp Ala Cys Ser Ser Lys Ala Val Val Ser Leu Arg 225 230 235 240 Cys Leu Ala Cys Gly Val Asn Leu Asn Ser Ser Arg Gln Ser Arg Ile 245 250 255 Val Gly Gly Glu Ser Ala Leu Pro Gly Ala Trp Pro Trp Gln Val Ser 260 265 270 Leu His Val Gln Asn Val His Val Cys Gly Gly Ser Ile Ile Thr Pro 275 280 285 Glu Trp Ile Val Thr Ala Ala His Cys Val Glu Lys Pro Leu Asn Asn 290 295 300 Pro Trp His Trp Thr Ala Phe Ala Gly Ile Leu Arg Gln Ser Phe Met 305 310 315 320 Phe Tyr Gly Ala Gly Tyr Gln Val Gln Lys Val Ile Ser His Pro Asn 325 330 335 Tyr Asp Ser Lys Thr Lys Asn Asn Asp Ile Ala Leu Met Lys Leu Gln 340 345 350 Lys Pro Leu Thr Phe Asn Asp Leu Val Lys Pro Val Cys Leu Pro Asn 355 360 365 Pro Gly Met Met Leu Gln Pro Glu Gln Leu Cys Trp Ile Ser Gly Trp 370 375 380 Gly Ala Thr Glu Glu Lys Gly Lys Thr Ser Glu Val Leu Asn Ala Ala 385 390 395 400 Lys Val Leu Leu Ile Glu Thr Gln Arg Cys Asn Ser Arg Tyr Val Tyr 405 410 415 Asp Asn Leu Ile Thr Pro Ala Met Ile Cys Ala Gly Phe Leu Gln Gly 420 425 430 Asn Val Asp Ser Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Thr Ser 435 440 445 Asn Asn Asn Ile Trp Trp Leu Ile Gly Asp Thr Ser Trp Gly Ser Gly 450 455 460 Cys Ala Lys Ala Tyr Arg Pro Gly Val Tyr Gly Asn Val Met Val Phe 465 470 475 480 Thr Asp Trp Ile Tyr Arg Gln Met Lys Ala Asn Gly 485 490 <210> SEQ ID NO 38 <211> LENGTH: 2079 <212> TYPE: DNA <213> ORGANISM: Homo sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (251)...(1522) <223> OTHER INFORMATION: Nucleotide sequence encoding transmembrane protease, serine 4 (TMPRSS4) <300> PUBLICATION INFORMATION: <308> DATABASE ACCESSION NUMBER: GenBank NM016425 <309> DATABASE ENTRY DATE: 2000-11-06 <400> SEQUENCE: 38 gagaggcagc agcttgttca gcggacaagg atgctgggcg tgagggacca aggcctgccc 60 tgcactcggg cctcctccag ccagtgctga ccagggactt ctgacctgct ggccagccag 120 gacctgtgtg gggaggccct cctgctgcct tggggtgaca atctcagctc caggctacag 180 ggagaccggg aggatcacag agccagcatg gtacaggatc ctgacagtga tcaacctctg 240 aacagcctcg atg tca aac ccc tgc gca aac ccc gta tcc cca tgg aga 289 Met Ser Asn Pro Cys Ala Asn Pro Val Ser Pro Trp Arg 1 5 10 cct tca gaa agt gtg ggg atc ccc atc atc ata gca cta ctg agc ctg 337Pro Ser Glu Ser Val Gly Ile Pro Ile Ile Ile Ala Leu Leu Ser Leu 15 20 25 gcg agt atc atc att gtg gtt gtc ctc atc aag gtg att ctg gat aaa 385 Ala Ser Ile Ile Ile Val Val Val Leu Ile Lys Val Ile Leu Asp Lys 30 35 40 45 tac tac ttc ctc tgc ggg cag cct ctc cac ttc atc ccg agg aag cag 433Tyr Tyr Phe Leu Cys Gly Gln Pro Leu His Phe Ile Pro Arg Lys Gln 50 55 60 ctg tgt gac gga gag ctg gac tgt ccc ttg ggg gag gac gag gag cac 481Leu Cys Asp Gly Glu Leu Asp Cys Pro Leu Gly Glu Asp Glu Glu His 65 70 75 tgt gtc aag agc ttc ccc gaa ggg cct gca gtg gca gtc cgc ctc tcc 529Cys Val Lys Ser Phe Pro Glu Gly Pro Ala Val Ala Val Arg Leu Ser 80 85 90 aag gac cga tcc aca ctg cag gtg ctg gac tcg gcc aca ggg aac tgg 577 Lys Asp Arg Ser Thr Leu Gln Val Leu Asp Ser Ala Thr Gly Asn Trp 95 100 105 ttc tct gcc tgt ttc gac aac ttc aca gaa gct ctc gct gag aca gcc 625 Phe Ser Ala Cys Phe Asp Asn Phe Thr Glu Ala Leu Ala Glu Thr Ala 110 115 120 125 tgt agg cag atg ggc tac agc agc aaa ccc act ttc aga gct gtg gag 673 Cys Arg Gln Met Gly Tyr Ser Ser Lys Pro Thr Phe Arg Ala Val Glu 130 135 140 att ggc cca gac cag gat ctg gat gtt gtt gaa atc aca gaa aac agc 721 Ile Gly Pro Asp Gln Asp Leu Asp Val Val Glu Ile Thr Glu Asn Ser 145 150 155 cag gag ctt cgc atg cgg aac tca agt ggg ccc tgt ctc tca ggc tcc 769 Gln Glu Leu Arg Met Arg Asn Ser Ser Gly Pro Cys Leu Ser Gly Ser 160 165 170 ctg gtc tcc ctg cac tgt ctt gcc tgt ggg aag agc ctg aag acc ccc 817 Leu Val Ser Leu His Cys Leu Ala Cys Gly Lys Ser Leu Lys Thr Pro 175 180 185 cgt gtg gtg ggt ggg gag gag gcc tct gtg gat tct tgg cct tgg cag 865 Arg Val Val Gly Gly Glu Glu Ala Ser Val Asp Ser Trp Pro Trp Gln 190 195 200 205 gtc agc atc cag tac gac aaa cag cac gtc tgt gga ggg agc atc ctg 913 Val Ser Ile Gln Tyr Asp Lys Gln His Val Cys Gly Gly Ser Ile Leu 210 215 220 gac ccc cac tgg gtc ctc acg gca gcc cac tgc ttc agg aaa cat acc 961 Asp Pro His Trp Val Leu Thr Ala Ala His Cys Phe Arg Lys His Thr 225 230 235 gat gtg ttc aac tgg aag gtg cgg gca ggc tca gac aaa ctg ggc agc 1009 Asp Val Phe Asn Trp Lys Val Arg Ala Gly Ser Asp Lys Leu Gly Ser 240 245 250 ttc cca tcc ctg gct gtg gcc aag atc atc atc att gaa ttc aac ccc 1057 Phe Pro Ser Leu Ala Val Ala Lys Ile Ile Ile Ile Glu Phe Asn Pro 255 260 265 atg tac ccc aaa gac aat gac atc gcc ctc atg aag ctg cag ttc cca 1105 Met Tyr Pro Lys Asp Asn Asp Ile Ala Leu Met Lys Leu Gln Phe Pro 270 275 280 285 ctc act ttc tca ggc aca gtc agg ccc atc tgt ctg ccc ttc ttt gat 1153 Leu Thr Phe Ser Gly Thr Val Arg Pro Ile Cys Leu Pro Phe Phe Asp 290 295 300 gag gag ctc act cca gcc acc cca ctc tgg atc att gga tgg ggc ttt 1201 Glu Glu Leu Thr Pro Ala Thr Pro Leu Trp Ile Ile Gly Trp Gly Phe 305 310 315 acg aag cag aat gga ggg aag atg tct gac ata ctg ctg cag gcg tca 1249 Thr Lys Gln Asn Gly Gly Lys Met Ser Asp Ile Leu Leu Gln Ala Ser 320 325 330 gtc cag gtc att gac agc aca cgg tgc aat gca gac gat gcg tac cag 1297 Val Gln Val Ile Asp Ser Thr Arg Cys Asn Ala Asp Asp Ala Tyr Gln 335 340 345 ggg gaa gtc acc gag aag atg atg tgt gca ggc atc ccg gaa ggg ggt 1345 Gly Glu Val Thr Glu Lys Met Met Cys Ala Gly Ile Pro Glu Gly Gly 350 355 360 365 gtg gac acc tgc cag ggt gac agt ggt ggg ccc ctg atg tac caa tct 1393 Val Asp Thr Cys Gln Gly Asp Ser Gly Gly Pro Leu Met Tyr Gln Ser 370 375 380 gac cag tgg cat gtg gtg ggc atc gtt agc tgg ggc tat ggc tgc ggg 1441 Asp Gln Trp His Val Val Gly Ile Val Ser Trp Gly Tyr Gly Cys Gly 385 390 395 ggc ccg agc acc cca gga gta tac acc aag gtc tca gcc tat ctc aac 1489 Gly Pro Ser Thr Pro Gly Val Tyr Thr Lys Val Ser Ala Tyr Leu Asn 400 405 410 tgg atc tac aat gtc tgg aag gct gag ctg taa tgctgctgcc cctttgcagt 1542 Trp Ile Tyr Asn Val Trp Lys Ala Glu Leu * 415 420 gctgggagcc gcttccttcc tgccctgccc acctggggat cccccaaagt cagacacaga 1602 gcaagagtcc ccttgggtac acccctctgc ccacagcctc agcatttctt ggagcagcaa 1662 agggcctcaa ttcctgtaag agaccctcgc agcccagagg cgcccagagg aagtcagcag 1722 ccctagctcg gccacacttg gtgctcccag catcccaggg agagacacag cccactgaac 1782 aaggtctcag gggtattgct aagccaagaa ggaactttcc cacactactg aatggaagca 1842 ggctgtcttg taaaagccca gatcactgtg ggctggagag gagaaggaaa gggtctgcgc 1902 cagccctgtc cgtcttcacc catccccaag cctactagag caagaaacca gttgtaatat 1962 aaaatgcact gccctactgt tggtatgact accgttacct actgttgtca ttgttattac 2022 agctatggcc actattatta aagagctgtg taacatcaaa aaaaaaaaaa aaaaaaa 2079 <210> SEQ ID NO 39 <211> LENGTH: 423 <212> TYPE: PRT <213> ORGANISM: Homo sapien <400> SEQUENCE: 39 Met Ser Asn Pro Cys Ala Asn Pro Val Ser Pro Trp Arg Pro Ser Glu 1 5 10 15 Ser Val Gly Ile Pro Ile Ile Ile Ala Leu Leu Ser Leu Ala Ser Ile 20 25 30 Ile Ile Val Val Val Leu Ile Lys Val Ile Leu Asp Lys Tyr Tyr Phe 35 40 45 Leu Cys Gly Gln Pro Leu His Phe Ile Pro Arg Lys Gln Leu Cys Asp 50 55 60 Gly Glu Leu Asp Cys Pro Leu Gly Glu Asp Glu Glu His Cys Val Lys 65 70 75 80 Ser Phe Pro Glu Gly Pro Ala Val Ala Val Arg Leu Ser Lys Asp Arg 85 90 95 Ser Thr Leu Gln Val Leu Asp Ser Ala Thr Gly Asn Trp Phe Ser Ala 100 105 110 Cys Phe Asp Asn Phe Thr Glu Ala Leu Ala Glu Thr Ala Cys Arg Gln 115 120 125 Met Gly Tyr Ser Ser Lys Pro Thr Phe Arg Ala Val Glu Ile Gly Pro 130 135 140 Asp Gln Asp Leu Asp Val Val Glu Ile Thr Glu Asn Ser Gln Glu Leu 145 150 155 160 Arg Met Arg Asn Ser Ser Gly Pro Cys Leu Ser Gly Ser Leu Val Ser 165 170 175 Leu His Cys Leu Ala Cys Gly Lys Ser Leu Lys Thr Pro Arg Val Val 180 185 190 Gly Gly Glu Glu Ala Ser Val Asp Ser Trp Pro Trp Gln Val Ser Ile 195 200 205 Gln Tyr Asp Lys Gln His Val Cys Gly Gly Ser Ile Leu Asp Pro His 210 215 220 Trp Val Leu Thr Ala Ala His Cys Phe Arg Lys His Thr Asp Val Phe 225 230 235 240 Asn Trp Lys Val Arg Ala Gly Ser Asp Lys Leu Gly Ser Phe Pro Ser 245 250 255 Leu Ala Val Ala Lys Ile Ile Ile Ile Glu Phe Asn Pro Met Tyr Pro 260 265 270 Lys Asp Asn Asp Ile Ala Leu Met Lys Leu Gln Phe Pro Leu Thr Phe 275 280 285 Ser Gly Thr Val Arg Pro Ile Cys Leu Pro Phe Phe Asp Glu Glu Leu 290 295 300 Thr Pro Ala Thr Pro Leu Trp Ile Ile Gly Trp Gly Phe Thr Lys Gln 305 310 315 320 Asn Gly Gly Lys Met Ser Asp Ile Leu Leu Gln Ala Ser Val Gln Val 325 330 335 Ile Asp Ser Thr Arg Cys Asn Ala Asp Asp Ala Tyr Gln Gly Glu Val 340 345 350 Thr Glu Lys Met Met Cys Ala Gly Ile Pro Glu Gly Gly Val Asp Thr 355 360 365 Cys Gln Gly Asp Ser Gly Gly Pro Leu Met Tyr Gln Ser Asp Gln Trp 370 375 380 His Val Val Gly Ile Val Ser Trp Gly Tyr Gly Cys Gly Gly Pro Ser 385 390 395 400 Thr Pro Gly Val Tyr Thr Lys Val Ser Ala Tyr Leu Asn Trp Ile Tyr 405 410 415 Asn Val Trp Lys Ala Glu Leu 420 <210> SEQ ID NO 40 <211> LENGTH: 1471 <212> TYPE: DNA <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: DESC1 gene <220> FEATURE: <221> NAME/KEY: misc_feature <222> LOCATION: (626)...(1324) <223> OTHER INFORMATION: protease domain <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (56)...(1324) <400> SEQUENCE: 40 tgacttggat gtagacctcg accttcacag gactcttcat tgctggttgg caatg atg 58 Met 1 tat cgg cca gat gtg gtg agg gct agg aaa aga gtt tgt tgg gaa ccc 106Tyr Arg Pro Asp Val Val Arg Ala Arg Lys Arg Val Cys Trp Glu Pro 5 10 15 tgg gtt atc ggc ctc gtc ats ttc ata tcc ctg att gtc ctg gca gtg 154 Trp Val Ile Gly Leu Val Xaa Phe Ile Ser Leu Ile Val Leu Ala Val 20 25 30 tgc att gga stc act gtt cat tat gtg aga tat aat caa aag aag acc 202 Cys Ile Gly Xaa Thr Val His Tyr Val Arg Tyr Asn Gln Lys Lys Thr 35 40 45 tac aat tac tat agc aca ttg tca ttt aca act gac aaa cta tat gct 250 Tyr Asn Tyr Tyr Ser Thr Leu Ser Phe Thr Thr Asp Lys Leu Tyr Ala 50 55 60 65 gag ttt ggc aga gag gct tct aac aat ttt aca gaa atg agc cag aga 298 Glu Phe Gly Arg Glu Ala Ser Asn Asn Phe Thr Glu Met Ser Gln Arg 70 75 80 ctt gaa tca atg gtg aaa aat gca ttt tat aaa tct cca tta agg gaa 346 Leu Glu Ser Met Val Lys Asn Ala Phe Tyr Lys Ser Pro Leu Arg Glu 85 90 95 gaa ttt gtc aag tct cag gtt atc aag ttc agt caa cag aag cat gga 394Glu Phe Val Lys Ser Gln Val Ile Lys Phe Ser Gln Gln Lys His Gly 100 105 110 gtg ttg gct cat atg ctg ttg att tgt aga ttt cac tct act gag gat 442 Val Leu Ala His Met Leu Leu Ile Cys Arg Phe His Ser Thr Glu Asp 115 120 125 cct gaa act gta gat aaa att gtt caa ctt gtt tta cat gaa aag ctg 490Pro Glu Thr Val Asp Lys Ile Val Gln Leu Val Leu His Glu Lys Leu 130 135 140 145 caa gat gct gta gga ccc cct aaa gta gat cct cac tca gtt aaa att 538 Gln Asp Ala Val Gly Pro Pro Lys Val Asp Pro His Ser Val Lys Ile 150 155 160 aaa aaa atc aac aag aca gaa aca gac agc tat cta aac cat tgc tgc 586 Lys Lys Ile Asn Lys Thr Glu Thr Asp Ser Tyr Leu Asn His Cys Cys 165 170 175 gga aca cga aga agt aaa act cta ggt cag agt ctc agg atc gtt ggt 634 Gly Thr Arg Arg Ser Lys Thr Leu Gly Gln Ser Leu Arg Ile Val Gly 180 185 190 ggg aca gaa gta gaa gag ggt gaa tgg ccc tgg cag gct agc ctg cag 682 Gly Thr Glu Val Glu Glu Gly Glu Trp Pro Trp Gln Ala Ser Leu Gln 195 200 205 tgg gat ggg agt cat cgc tgt gga gca acc tta att aat gcc aca tgg 730 Trp Asp Gly Ser His Arg Cys Gly Ala Thr Leu Ile Asn Ala Thr Trp 210 215 220 225 ctt gtg agt gct gct cac tgt ttt aca aca tat aag aac cct gcc aga 778 Leu Val Ser Ala Ala His Cys Phe Thr Thr Tyr Lys Asn Pro Ala Arg 230 235 240 tgg act gct tcc ttt gga gta aca ata aaa cct tcg aaa atg aaa cgg 826 Trp Thr Ala Ser Phe Gly Val Thr Ile Lys Pro Ser Lys Met Lys Arg 245 250 255 ggt ctc cgg aga ata att gtc cat gaa aaa tac aaa cac cca tca cat 874 Gly Leu Arg Arg Ile Ile Val His Glu Lys Tyr Lys His Pro Ser His 260 265 270 gac tat gat att tct ctt gca gag ctt tct agc cct gtt ccc tac aca 922 Asp Tyr Asp Ile Ser Leu Ala Glu Leu Ser Ser Pro Val Pro Tyr Thr 275 280 285 aat gca gta cat aga gtt tgt ctc cct gat gca tcc tat gag ttt caa 970 Asn Ala Val His Arg Val Cys Leu Pro Asp Ala Ser Tyr Glu Phe Gln 290 295 300 305 cca ggt gat gtg atg ttt gtg aca gga ttt gga gca ctg aaa aat gat 1018 Pro Gly Asp Val Met Phe Val Thr Gly Phe Gly Ala Leu Lys Asn Asp 310 315 320 ggt tac agt caa aat cat ctt cga caa gca cag gtg act ctc ata gac 1066 Gly Tyr Ser Gln Asn His Leu Arg Gln Ala Gln Val Thr Leu Ile Asp 325 330 335 gct aca act tgc aat gaa cct caa gct tac aat gac gcc ata act cct 1114 Ala Thr Thr Cys Asn Glu Pro Gln Ala Tyr Asn Asp Ala Ile Thr Pro 340 345 350 aga atg tta tgt gct ggc tcc tta gaa gga aaa aca gat gca tgc cag 1162 Arg Met Leu Cys Ala Gly Ser Leu Glu Gly Lys Thr Asp Ala Cys Gln 355 360 365 ggt gac tct gga gga cca ctg gtt agt tca gat gct aga gat atc tgg 1210 Gly Asp Ser Gly Gly Pro Leu Val Ser Ser Asp Ala Arg Asp Ile Trp 370 375 380 385 tac ctt gct gga ata gtg agc tsg gga gat gaa tgt gcg aaa ccc aac 1258 Tyr Leu Ala Gly Ile Val Ser Xaa Gly Asp Glu Cys Ala Lys Pro Asn 390 395 400 aag cct ggt gtt tat act aga gtt acg gcc ttg cgg gac tgg att act 1306 Lys Pro Gly Val Tyr Thr Arg Val Thr Ala Leu Arg Asp Trp Ile Thr 405 410 415 tca aaa act ggt atc taa gagagaaaag cctcatggaa cagataacat 1354 Ser Lys Thr Gly Ile * 420 ttttttttgt tttttgggtg tggaggccat ttttagagat acagaattgg agaagacttg 1414 caaaacagct agatttgact gatctcaata aactgtttgc ttgatgcaaa aaaaaaa 1471 <210> SEQ ID NO 41 <211> LENGTH: 422 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: VARIANT <222> LOCATION: 24 <223> OTHER INFORMATION: Xaa is Ile or Met <220> FEATURE: <221> NAME/KEY: VARIANT <222> LOCATION: 37 <223> OTHER INFORMATION: Xaa is Leu or Val <220> FEATURE: <221> NAME/KEY: VARIANT <222> LOCATION: 393 <223> OTHER INFORMATION: Xaa is Ser or Trp <400> SEQUENCE: 41 Met Tyr Arg Pro Asp Val Val Arg Ala Arg Lys Arg Val Cys Trp Glu 1 5 10 15 Pro Trp Val Ile Gly Leu Val Xaa Phe Ile Ser Leu Ile Val Leu Ala 20 25 30 Val Cys Ile Gly Xaa Thr Val His Tyr Val Arg Tyr Asn Gln Lys Lys 35 40 45 Thr Tyr Asn Tyr Tyr Ser Thr Leu Ser Phe Thr Thr Asp Lys Leu Tyr 50 55 60 Ala Glu Phe Gly Arg Glu Ala Ser Asn Asn Phe Thr Glu Met Ser Gln 65 70 75 80 Arg Leu Glu Ser Met Val Lys Asn Ala Phe Tyr Lys Ser Pro Leu Arg 85 90 95 Glu Glu Phe Val Lys Ser Gln Val Ile Lys Phe Ser Gln Gln Lys His 100 105 110 Gly Val Leu Ala His Met Leu Leu Ile Cys Arg Phe His Ser Thr Glu 115 120 125 Asp Pro Glu Thr Val Asp Lys Ile Val Gln Leu Val Leu His Glu Lys 130 135 140 Leu Gln Asp Ala Val Gly Pro Pro Lys Val Asp Pro His Ser Val Lys 145 150 155 160 Ile Lys Lys Ile Asn Lys Thr Glu Thr Asp Ser Tyr Leu Asn His Cys 165 170 175 Cys Gly Thr Arg Arg Ser Lys Thr Leu Gly Gln Ser Leu Arg Ile Val 180 185 190 Gly Gly Thr Glu Val Glu Glu Gly Glu Trp Pro Trp Gln Ala Ser Leu 195 200 205 Gln Trp Asp Gly Ser His Arg Cys Gly Ala Thr Leu Ile Asn Ala Thr 210 215 220 Trp Leu Val Ser Ala Ala His Cys Phe Thr Thr Tyr Lys Asn Pro Ala 225 230 235 240 Arg Trp Thr Ala Ser Phe Gly Val Thr Ile Lys Pro Ser Lys Met Lys 245 250 255 Arg Gly Leu Arg Arg Ile Ile Val His Glu Lys Tyr Lys His Pro Ser 260 265 270 His Asp Tyr Asp Ile Ser Leu Ala Glu Leu Ser Ser Pro Val Pro Tyr 275 280 285 Thr Asn Ala Val His Arg Val Cys Leu Pro Asp Ala Ser Tyr Glu Phe 290 295 300 Gln Pro Gly Asp Val Met Phe Val Thr Gly Phe Gly Ala Leu Lys Asn 305 310 315 320 Asp Gly Tyr Ser Gln Asn His Leu Arg Gln Ala Gln Val Thr Leu Ile 325 330 335 Asp Ala Thr Thr Cys Asn Glu Pro Gln Ala Tyr Asn Asp Ala Ile Thr 340 345 350 Pro Arg Met Leu Cys Ala Gly Ser Leu Glu Gly Lys Thr Asp Ala Cys 355 360 365 Gln Gly Asp Ser Gly Gly Pro Leu Val Ser Ser Asp Ala Arg Asp Ile 370 375 380 Trp Tyr Leu Ala Gly Ile Val Ser Xaa Gly Asp Glu Cys Ala Lys Pro 385 390 395 400 Asn Lys Pro Gly Val Tyr Thr Arg Val Thr Ala Leu Arg Asp Trp Ile 405 410 415 Thr Ser Lys Thr Gly Ile 420 <210> SEQ ID NO 42 <211> LENGTH: 1257 <212> TYPE: DNA <213> ORGANISM: Homo sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (1)...(1257) <223> OTHER INFORMATION: Nucleotide sequence encoding MTSP9 <400> SEQUENCE: 42 atg atg tat cgg aca gta gga ttt ggc acc cga agc aga aat ctg aag 48 Met Met Tyr Arg Thr Val Gly Phe Gly Thr Arg Ser Arg Asn Leu Lys 1 5 10 15 cca tgg atg att gcc gtt ctc att gtg ttg tcc ctg aca gtg gtg gca 96 Pro Trp Met Ile Ala Val Leu Ile Val Leu Ser Leu Thr Val Val Ala 20 25 30 gtg acc ata ggt ctc ctg gtt cac ttc cta gta ttt gac caa aaa aag 144 Val Thr Ile Gly Leu Leu Val His Phe Leu Val Phe Asp Gln Lys Lys 35 40 45 gag tac tat cat ggc tcc ttt aaa att tta gat cca caa atc aat aac 192 Glu Tyr Tyr His Gly Ser Phe Lys Ile Leu Asp Pro Gln Ile Asn Asn 50 55 60 aat ttc gga caa agc aac aca tat caa ctt aag gac tta cga gag acg 240 Asn Phe Gly Gln Ser Asn Thr Tyr Gln Leu Lys Asp Leu Arg Glu Thr 65 70 75 80 acc gaa aat ttg gtg gat gag ata ttt ata gat tca gcc tgg aag aaa 288 Thr Glu Asn Leu Val Asp Glu Ile Phe Ile Asp Ser Ala Trp Lys Lys 85 90 95 aat tat atc aag aac caa gta gtc aga ctg act cca gag gaa gat ggt 336Asn Tyr Ile Lys Asn Gln Val Val Arg Leu Thr Pro Glu Glu Asp Gly 100 105 110 gtg aaa gta gat gtc att atg gtg ttc cag ttc ccc tct act gaa caa 384 Val Lys Val Asp Val Ile Met Val Phe Gln Phe Pro Ser Thr Glu Gln 115 120 125 agg gca gta aga gag aag aaa atc caa agc atc tta aat cag aag ata 432 Arg Ala Val Arg Glu Lys Lys Ile Gln Ser Ile Leu Asn Gln Lys Ile 130 135 140 agg aat tta aga gcc ttg cca ata aat gcc tca tca gtt caa gtt aat 480 Arg Asn Leu Arg Ala Leu Pro Ile Asn Ala Ser Ser Val Gln Val Asn 145 150 155 160 gca atg agc tca tca aca ggg gag tta act gtc caa gca agt tgt ggt 528Ala Met Ser Ser Ser Thr Gly Glu Leu Thr Val Gln Ala Ser Cys Gly 165 170 175 aaa cga gtt gtt cca tta aac gtc aac aga ata gca tct gga gtc att 576 Lys Arg Val Val Pro Leu Asn Val Asn Arg Ile Ala Ser Gly Val Ile 180 185 190 gca ccc aag gcg gcc tgg cct tgg caa gct tcc ctt cag tat gat aac 624 Ala Pro Lys Ala Ala Trp Pro Trp Gln Ala Ser Leu Gln Tyr Asp Asn 195 200 205 atc cat cag tgt ggg gcc acc ttg att agt aac aca tgg ctt gtc act 672 Ile His Gln Cys Gly Ala Thr Leu Ile Ser Asn Thr Trp Leu Val Thr 210 215 220 gca gca cac tgc ttc cag aag tat aaa aat cca cat caa tgg act gtt 720 Ala Ala His Cys Phe Gln Lys Tyr Lys Asn Pro His Gln Trp Thr Val 225 230 235 240 agt ttt gga aca aaa atc aac cct ccc tta atg aaa aga aat gtc aga 768 Ser Phe Gly Thr Lys Ile Asn Pro Pro Leu Met Lys Arg Asn Val Arg 245 250 255 aga ttt att atc cat gag aag tac cgc tct gca gca aga gag tac gac 816 Arg Phe Ile Ile His Glu Lys Tyr Arg Ser Ala Ala Arg Glu Tyr Asp 260 265 270 att gct gtt gtg cag gtc tct tcc aga gtc acc ttt tcg gat gac ata 864 Ile Ala Val Val Gln Val Ser Ser Arg Val Thr Phe Ser Asp Asp Ile 275 280 285 cgc cgg att tgt ttg cca gaa gcc tct gca tcc ttc caa cca aat ttg 912 Arg Arg Ile Cys Leu Pro Glu Ala Ser Ala Ser Phe Gln Pro Asn Leu 290 295 300 act gtc cac atc aca gga ttt gga gca ctt tac tat ggt ggg gaa tcc 960 Thr Val His Ile Thr Gly Phe Gly Ala Leu Tyr Tyr Gly Gly Glu Ser 305 310 315 320 caa aat gat ctc cga gaa gcc aga gtg aaa atc ata agt gac gat gtc 1008 Gln Asn Asp Leu Arg Glu Ala Arg Val Lys Ile Ile Ser Asp Asp Val 325 330 335 tgc aag caa cca cag gtg tat ggc aat gat ata aaa cct gga atg ttc 1056 Cys Lys Gln Pro Gln Val Tyr Gly Asn Asp Ile Lys Pro Gly Met Phe 340 345 350 tgt gcc gga tat atg gaa gga att tat gat gcc tgc agg ggt gat tct 1104 Cys Ala Gly Tyr Met Glu Gly Ile Tyr Asp Ala Cys Arg Gly Asp Ser 355 360 365 ggg gga cct tta gtc aca agg gat ctg aaa gat acg tgg tat ctc att 1152 Gly Gly Pro Leu Val Thr Arg Asp Leu Lys Asp Thr Trp Tyr Leu Ile 370 375 380 gga att gta agc tgg gga gat aac tgt ggt caa aag gac aag cct gga 1200 Gly Ile Val Ser Trp Gly Asp Asn Cys Gly Gln Lys Asp Lys Pro Gly 385 390 395 400 gtc tac aca caa gtg act tat tac cga aac tgg att gct tca aaa aca 1248 Val Tyr Thr Gln Val Thr Tyr Tyr Arg Asn Trp Ile Ala Ser Lys Thr 405 410 415 ggc atc taa 1257 Gly Ile * <210> SEQ ID NO 43 <211> LENGTH: 418 <212> TYPE: PRT <213> ORGANISM: Homo sapien <400> SEQUENCE: 43 Met Met Tyr Arg Thr Val Gly Phe Gly Thr Arg Ser Arg Asn Leu Lys 1 5 10 15 Pro Trp Met Ile Ala Val Leu Ile Val Leu Ser Leu Thr Val Val Ala 20 25 30 Val Thr Ile Gly Leu Leu Val His Phe Leu Val Phe Asp Gln Lys Lys 35 40 45 Glu Tyr Tyr His Gly Ser Phe Lys Ile Leu Asp Pro Gln Ile Asn Asn 50 55 60 Asn Phe Gly Gln Ser Asn Thr Tyr Gln Leu Lys Asp Leu Arg Glu Thr 65 70 75 80 Thr Glu Asn Leu Val Asp Glu Ile Phe Ile Asp Ser Ala Trp Lys Lys 85 90 95 Asn Tyr Ile Lys Asn Gln Val Val Arg Leu Thr Pro Glu Glu Asp Gly 100 105 110 Val Lys Val Asp Val Ile Met Val Phe Gln Phe Pro Ser Thr Glu Gln 115 120 125 Arg Ala Val Arg Glu Lys Lys Ile Gln Ser Ile Leu Asn Gln Lys Ile 130 135 140 Arg Asn Leu Arg Ala Leu Pro Ile Asn Ala Ser Ser Val Gln Val Asn 145 150 155 160 Ala Met Ser Ser Ser Thr Gly Glu Leu Thr Val Gln Ala Ser Cys Gly 165 170 175 Lys Arg Val Val Pro Leu Asn Val Asn Arg Ile Ala Ser Gly Val Ile 180 185 190 Ala Pro Lys Ala Ala Trp Pro Trp Gln Ala Ser Leu Gln Tyr Asp Asn 195 200 205 Ile His Gln Cys Gly Ala Thr Leu Ile Ser Asn Thr Trp Leu Val Thr 210 215 220 Ala Ala His Cys Phe Gln Lys Tyr Lys Asn Pro His Gln Trp Thr Val 225 230 235 240 Ser Phe Gly Thr Lys Ile Asn Pro Pro Leu Met Lys Arg Asn Val Arg 245 250 255 Arg Phe Ile Ile His Glu Lys Tyr Arg Ser Ala Ala Arg Glu Tyr Asp 260 265 270 Ile Ala Val Val Gln Val Ser Ser Arg Val Thr Phe Ser Asp Asp Ile 275 280 285 Arg Arg Ile Cys Leu Pro Glu Ala Ser Ala Ser Phe Gln Pro Asn Leu 290 295 300 Thr Val His Ile Thr Gly Phe Gly Ala Leu Tyr Tyr Gly Gly Glu Ser 305 310 315 320 Gln Asn Asp Leu Arg Glu Ala Arg Val Lys Ile Ile Ser Asp Asp Val 325 330 335 Cys Lys Gln Pro Gln Val Tyr Gly Asn Asp Ile Lys Pro Gly Met Phe 340 345 350 Cys Ala Gly Tyr Met Glu Gly Ile Tyr Asp Ala Cys Arg Gly Asp Ser 355 360 365 Gly Gly Pro Leu Val Thr Arg Asp Leu Lys Asp Thr Trp Tyr Leu Ile 370 375 380 Gly Ile Val Ser Trp Gly Asp Asn Cys Gly Gln Lys Asp Lys Pro Gly 385 390 395 400 Val Tyr Thr Gln Val Thr Tyr Tyr Arg Asn Trp Ile Ala Ser Lys Thr 405 410 415 Gly Ile <210> SEQ ID NO 44 <211> LENGTH: 2130 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (1)...(2104) <223> OTHER INFORMATION: Nucleotide sequence encoding MTSP10, including MTSP10 protease domain <400> SEQUENCE: 44 ata aac ctg gtt tat aca aca tct gcc ttc tcc aaa ttt tat gag cag 48 Ile Asn Leu Val Tyr Thr Thr Ser Ala Phe Ser Lys Phe Tyr Glu Gln 1 5 10 15 tct gtt gtt gca gat gtc agc agc aac aac aaa ggc ggc ctc ctt gtc 96 Ser Val Val Ala Asp Val Ser Ser Asn Asn Lys Gly Gly Leu Leu Val 20 25 30 cac ttt tgg att gtt ttt gtc atg cca cgt gcc aaa ggc cac atc ttc 144 His Phe Trp Ile Val Phe Val Met Pro Arg Ala Lys Gly His Ile Phe 35 40 45 tgt gaa gac tgt gtt gcc gcc atc ttg aag gac tcc atc cag aca agc 192 Cys Glu Asp Cys Val Ala Ala Ile Leu Lys Asp Ser Ile Gln Thr Ser 50 55 60 atc ata aac cgg acc tct gtg ggg agc ttg cag gga ctg gct gtg gac 240 Ile Ile Asn Arg Thr Ser Val Gly Ser Leu Gln Gly Leu Ala Val Asp 65 70 75 80 atg gac tct gtg gta cta aat gct ggg ctt cgg tca gat tac tcg tca 288Met Asp Ser Val Val Leu Asn Ala Gly Leu Arg Ser Asp Tyr Ser Ser 85 90 95 acc ata gga tct gac aaa ggc tgc tct cag tac ttc tat gca gag cat 336Thr Ile Gly Ser Asp Lys Gly Cys Ser Gln Tyr Phe Tyr Ala Glu His 100 105 110 ctg tct ctc cac tac ccg ctg gag att tct gca gcc tca ggg agg ctg 384Leu Ser Leu His Tyr Pro Leu Glu Ile Ser Ala Ala Ser Gly Arg Leu 115 120 125 atg tgt cac ttc aag ctg gtg gcc ata gtg ggc tac ctg att cgt ctc 432Met Cys His Phe Lys Leu Val Ala Ile Val Gly Tyr Leu Ile Arg Leu 130 135 140 tca atc aag tcc atc caa atc gaa gcc gac aac tgt gtc act gac tcc 480Ser Ile Lys Ser Ile Gln Ile Glu Ala Asp Asn Cys Val Thr Asp Ser 145 150 155 160 ctg acc att tac gac tcc ctt ttg ccc atc cgg agc agc atc ttg tac 528 Leu Thr Ile Tyr Asp Ser Leu Leu Pro Ile Arg Ser Ser Ile Leu Tyr 165 170 175 aga att tgt gaa ccc aca aga aca tta atg tca ttt gtt tct aca aat 576 Arg Ile Cys Glu Pro Thr Arg Thr Leu Met Ser Phe Val Ser Thr Asn 180 185 190 aat ctc atg ttg gtg aca ttt aag tct cct cat ata cgg agg ctc tca 624 Asn Leu Met Leu Val Thr Phe Lys Ser Pro His Ile Arg Arg Leu Ser 195 200 205 gga atc cgg gca tat ttt gag gtc att cca gaa caa aag tgt gaa aac 672 Gly Ile Arg Ala Tyr Phe Glu Val Ile Pro Glu Gln Lys Cys Glu Asn 210 215 220 aca gtg ttg gtc aaa gac atc act ggc ttt gaa ggg aaa att tca agc 720 Thr Val Leu Val Lys Asp Ile Thr Gly Phe Glu Gly Lys Ile Ser Ser 225 230 235 240 cca tat tac ccg agc tac tat cct cca aaa tgc aag tgt acc tgg aaa 768 Pro Tyr Tyr Pro Ser Tyr Tyr Pro Pro Lys Cys Lys Cys Thr Trp Lys 245 250 255 ttt cag act tct cta tca act ctt ggc ata gca ctg aaa ttc tat aac 816 Phe Gln Thr Ser Leu Ser Thr Leu Gly Ile Ala Leu Lys Phe Tyr Asn 260 265 270 tat tca ata acc aag aag agt atg aaa ggc tgt gag cat gga tgg tgg 864 Tyr Ser Ile Thr Lys Lys Ser Met Lys Gly Cys Glu His Gly Trp Trp 275 280 285 gaa att tat gag cac atg tac tgt ggc tcc tac atg gat cat cag aca 912 Glu Ile Tyr Glu His Met Tyr Cys Gly Ser Tyr Met Asp His Gln Thr 290 295 300 att ttt cga gtg ccc agc cct ctg gtt cac att cag ctc cag tgc agt 960 Ile Phe Arg Val Pro Ser Pro Leu Val His Ile Gln Leu Gln Cys Ser 305 310 315 320 tca agg ctt tca ggc aag cca ctt ttg gca gaa tat ggc agt tac aac 1008 Ser Arg Leu Ser Gly Lys Pro Leu Leu Ala Glu Tyr Gly Ser Tyr Asn 325 330 335 atc agt caa ccc tgc cct gtg gga tct ttt aga tgc tcc tcc ggt tta 1056 Ile Ser Gln Pro Cys Pro Val Gly Ser Phe Arg Cys Ser Ser Gly Leu 340 345 350 tgt gtc cct cag gcc cag cgt ggt gat gga gta aat gac tgc ttt gat 1104 Cys Val Pro Gln Ala Gln Arg Gly Asp Gly Val Asn Asp Cys Phe Asp 355 360 365 gaa agt gat gaa ctg ttt tgc gtg agc cct caa cct gcc tgc aat acc 1152 Glu Ser Asp Glu Leu Phe Cys Val Ser Pro Gln Pro Ala Cys Asn Thr 370 375 380 agc tcc ttc agg cag cat ggc cct ctc atc tgt gat ggc ttc agg gac 1200 Ser Ser Phe Arg Gln His Gly Pro Leu Ile Cys Asp Gly Phe Arg Asp 385 390 395 400 tgt gag aat ggc cgg gat gag caa aac tgc act caa agt att cca tgc 1248 Cys Glu Asn Gly Arg Asp Glu Gln Asn Cys Thr Gln Ser Ile Pro Cys 405 410 415 aac aac aga act ttt aag tgt ggc aat gat att tgc ttt agg aaa caa 1296 Asn Asn Arg Thr Phe Lys Cys Gly Asn Asp Ile Cys Phe Arg Lys Gln 420 425 430 aat gca aaa tgt gat ggg aca gtg gat tgt cca gat gga agt gat gaa 1344 Asn Ala Lys Cys Asp Gly Thr Val Asp Cys Pro Asp Gly Ser Asp Glu 435 440 445 gaa ggc tgc acc tgc agc agg agt tcc tcc gcc ctt cac cgc atc atc 1392 Glu Gly Cys Thr Cys Ser Arg Ser Ser Ser Ala Leu His Arg Ile Ile 450 455 460 gga ggc aca gac acc ctg gag ggg ggt tgg ccg tgg cag gtc agc ctc 1440 Gly Gly Thr Asp Thr Leu Glu Gly Gly Trp Pro Trp Gln Val Ser Leu 465 470 475 480 cac ttt gtt gga tct gcc tac tgt ggt gcc tca gtc atc tcc agg gag 1488 His Phe Val Gly Ser Ala Tyr Cys Gly Ala Ser Val Ile Ser Arg Glu 485 490 495 tgg ctt ctt tct gca gcc cac tgt ttt cat gga aac agg ctg tca gat 1536 Trp Leu Leu Ser Ala Ala His Cys Phe His Gly Asn Arg Leu Ser Asp 500 505 510 ccc aca cca tgg act gca cac ctc ggg atg tat gtt cag ggg aat gcc 1584 Pro Thr Pro Trp Thr Ala His Leu Gly Met Tyr Val Gln Gly Asn Ala 515 520 525 aag ttt gtc tcc ccg gtg aga aga att gtg gtc cac gag tac tat aac 1632 Lys Phe Val Ser Pro Val Arg Arg Ile Val Val His Glu Tyr Tyr Asn 530 535 540 agt cag act ttt gat tat gat att gct ttg cta cag ctc agt att gcc 1680 Ser Gln Thr Phe Asp Tyr Asp Ile Ala Leu Leu Gln Leu Ser Ile Ala 545 550 555 560 tgg cct gag acc ctg aaa cag ctc att cag cca ata tgc att cct ccc 1728 Trp Pro Glu Thr Leu Lys Gln Leu Ile Gln Pro Ile Cys Ile Pro Pro 565 570 575 act ggt cag aga gtt cgc agt ggg gag aag tgc tgg gta act ggc tgg 1776 Thr Gly Gln Arg Val Arg Ser Gly Glu Lys Cys Trp Val Thr Gly Trp 580 585 590 ggg cga aga cac gaa gca gat aat aaa ggc tcc ctc gtt ctg cag caa 1824 Gly Arg Arg His Glu Ala Asp Asn Lys Gly Ser Leu Val Leu Gln Gln 595 600 605 gcg gag gta gag ctc att gat caa acg ctc tgt gtt tcc acc tac ggg 1872 Ala Glu Val Glu Leu Ile Asp Gln Thr Leu Cys Val Ser Thr Tyr Gly 610 615 620 atc atc act tct cgg atg ctc tgt gca ggc ata atg tca ggc aag aga 1920 Ile Ile Thr Ser Arg Met Leu Cys Ala Gly Ile Met Ser Gly Lys Arg 625 630 635 640 gat gcc tgc aaa gga gat tcg ggt gga cct tta tct tgt cga aga aaa 1968 Asp Ala Cys Lys Gly Asp Ser Gly Gly Pro Leu Ser Cys Arg Arg Lys 645 650 655 agt gat gga aaa tgg att ttg act ggc att gtt agc tgg gga cat gga 2016 Ser Asp Gly Lys Trp Ile Leu Thr Gly Ile Val Ser Trp Gly His Gly 660 665 670 tgt gga cga cca aac ttt cct ggt gtt tac aca agg gtg tca aac ttt 2064 Cys Gly Arg Pro Asn Phe Pro Gly Val Tyr Thr Arg Val Ser Asn Phe 675 680 685 gtt ccc tgg att cat aaa tat gtc cct tct ctt ttg taa t tgcaaaaaaa 2114 Val Pro Trp Ile His Lys Tyr Val Pro Ser Leu Leu * 690 695 700 aaaaaaaaaa aaaaaa 2130 <210> SEQ ID NO 45 <211> LENGTH: 700 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 45 Ile Asn Leu Val Tyr Thr Thr Ser Ala Phe Ser Lys Phe Tyr Glu Gln 1 5 10 15 Ser Val Val Ala Asp Val Ser Ser Asn Asn Lys Gly Gly Leu Leu Val 20 25 30 His Phe Trp Ile Val Phe Val Met Pro Arg Ala Lys Gly His Ile Phe 35 40 45 Cys Glu Asp Cys Val Ala Ala Ile Leu Lys Asp Ser Ile Gln Thr Ser 50 55 60 Ile Ile Asn Arg Thr Ser Val Gly Ser Leu Gln Gly Leu Ala Val Asp 65 70 75 80 Met Asp Ser Val Val Leu Asn Ala Gly Leu Arg Ser Asp Tyr Ser Ser 85 90 95 Thr Ile Gly Ser Asp Lys Gly Cys Ser Gln Tyr Phe Tyr Ala Glu His 100 105 110 Leu Ser Leu His Tyr Pro Leu Glu Ile Ser Ala Ala Ser Gly Arg Leu 115 120 125 Met Cys His Phe Lys Leu Val Ala Ile Val Gly Tyr Leu Ile Arg Leu 130 135 140 Ser Ile Lys Ser Ile Gln Ile Glu Ala Asp Asn Cys Val Thr Asp Ser 145 150 155 160 Leu Thr Ile Tyr Asp Ser Leu Leu Pro Ile Arg Ser Ser Ile Leu Tyr 165 170 175 Arg Ile Cys Glu Pro Thr Arg Thr Leu Met Ser Phe Val Ser Thr Asn 180 185 190 Asn Leu Met Leu Val Thr Phe Lys Ser Pro His Ile Arg Arg Leu Ser 195 200 205 Gly Ile Arg Ala Tyr Phe Glu Val Ile Pro Glu Gln Lys Cys Glu Asn 210 215 220 Thr Val Leu Val Lys Asp Ile Thr Gly Phe Glu Gly Lys Ile Ser Ser 225 230 235 240 Pro Tyr Tyr Pro Ser Tyr Tyr Pro Pro Lys Cys Lys Cys Thr Trp Lys 245 250 255 Phe Gln Thr Ser Leu Ser Thr Leu Gly Ile Ala Leu Lys Phe Tyr Asn 260 265 270 Tyr Ser Ile Thr Lys Lys Ser Met Lys Gly Cys Glu His Gly Trp Trp 275 280 285 Glu Ile Tyr Glu His Met Tyr Cys Gly Ser Tyr Met Asp His Gln Thr 290 295 300 Ile Phe Arg Val Pro Ser Pro Leu Val His Ile Gln Leu Gln Cys Ser 305 310 315 320 Ser Arg Leu Ser Gly Lys Pro Leu Leu Ala Glu Tyr Gly Ser Tyr Asn 325 330 335 Ile Ser Gln Pro Cys Pro Val Gly Ser Phe Arg Cys Ser Ser Gly Leu 340 345 350 Cys Val Pro Gln Ala Gln Arg Gly Asp Gly Val Asn Asp Cys Phe Asp 355 360 365 Glu Ser Asp Glu Leu Phe Cys Val Ser Pro Gln Pro Ala Cys Asn Thr 370 375 380 Ser Ser Phe Arg Gln His Gly Pro Leu Ile Cys Asp Gly Phe Arg Asp 385 390 395 400 Cys Glu Asn Gly Arg Asp Glu Gln Asn Cys Thr Gln Ser Ile Pro Cys 405 410 415 Asn Asn Arg Thr Phe Lys Cys Gly Asn Asp Ile Cys Phe Arg Lys Gln 420 425 430 Asn Ala Lys Cys Asp Gly Thr Val Asp Cys Pro Asp Gly Ser Asp Glu 435 440 445 Glu Gly Cys Thr Cys Ser Arg Ser Ser Ser Ala Leu His Arg Ile Ile 450 455 460 Gly Gly Thr Asp Thr Leu Glu Gly Gly Trp Pro Trp Gln Val Ser Leu 465 470 475 480 His Phe Val Gly Ser Ala Tyr Cys Gly Ala Ser Val Ile Ser Arg Glu 485 490 495 Trp Leu Leu Ser Ala Ala His Cys Phe His Gly Asn Arg Leu Ser Asp 500 505 510 Pro Thr Pro Trp Thr Ala His Leu Gly Met Tyr Val Gln Gly Asn Ala 515 520 525 Lys Phe Val Ser Pro Val Arg Arg Ile Val Val His Glu Tyr Tyr Asn 530 535 540 Ser Gln Thr Phe Asp Tyr Asp Ile Ala Leu Leu Gln Leu Ser Ile Ala 545 550 555 560 Trp Pro Glu Thr Leu Lys Gln Leu Ile Gln Pro Ile Cys Ile Pro Pro 565 570 575 Thr Gly Gln Arg Val Arg Ser Gly Glu Lys Cys Trp Val Thr Gly Trp 580 585 590 Gly Arg Arg His Glu Ala Asp Asn Lys Gly Ser Leu Val Leu Gln Gln 595 600 605 Ala Glu Val Glu Leu Ile Asp Gln Thr Leu Cys Val Ser Thr Tyr Gly 610 615 620 Ile Ile Thr Ser Arg Met Leu Cys Ala Gly Ile Met Ser Gly Lys Arg 625 630 635 640 Asp Ala Cys Lys Gly Asp Ser Gly Gly Pro Leu Ser Cys Arg Arg Lys 645 650 655 Ser Asp Gly Lys Trp Ile Leu Thr Gly Ile Val Ser Trp Gly His Gly 660 665 670 Cys Gly Arg Pro Asn Phe Pro Gly Val Tyr Thr Arg Val Ser Asn Phe 675 680 685 Val Pro Trp Ile His Lys Tyr Val Pro Ser Leu Leu 690 695 700 <210> SEQ ID NO 46 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 46 Leu Arg Ala Xaa Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 47 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Xaa is Alanine-therapeutic agent <400> SEQUENCE: 47 Leu Arg Ala Xaa Ala Arg Ala Xaa 1 5 <210> SEQ ID NO 48 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 48 Leu Arg Ser Xaa Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 49 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 49 Leu Arg Ser Xaa Ala Arg Ala Xaa 1 5 <210> SEQ ID NO 50 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 50 Leu Arg Pro Arg Phe Lys Ile Xaa 1 5 <210> SEQ ID NO 51 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 51 Arg Pro Arg Phe Lys Ile Xaa 1 5 <210> SEQ ID NO 52 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 52 Pro Arg Phe Lys Ile Xaa 1 5 <210> SEQ ID NO 53 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 53 Leu Arg Ser Lys Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 54 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 54 Arg Ser Lys Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 55 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 55 Ser Lys Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 56 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 56 Leu Arg Pro Arg Phe Arg Ile Xaa 1 5 <210> SEQ ID NO 57 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 57 Arg Pro Arg Phe Arg Ile Xaa 1 5 <210> SEQ ID NO 58 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 58 Pro Arg Phe Arg Ile Xaa 1 5 <210> SEQ ID NO 59 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 59 Leu Arg Ser Arg Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 60 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 60 Arg Ser Arg Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 61 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 61 Ser Arg Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 62 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 62 Leu Arg Ala Xaa Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 63 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 63 Leu Arg Ala Xaa Ala Arg Ala Xaa 1 5 <210> SEQ ID NO 64 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 64 Leu Arg Ser Xaa Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 65 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 65 Leu Arg Ser Xaa Ala Arg Ala Xaa 1 5 <210> SEQ ID NO 66 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 66 Leu Arg Pro Arg Phe Lys Ile Xaa 1 5 <210> SEQ ID NO 67 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 67 Arg Pro Arg Phe Lys Ile Xaa 1 5 <210> SEQ ID NO 68 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 68 Pro Arg Phe Lys Ile Xaa 1 5 <210> SEQ ID NO 69 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 69 Leu Arg Ser Lys Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 70 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 70 Arg Ser Lys Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 71 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (1)...(0) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 71 Ser Lys Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 72 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 72 Leu Arg Pro Arg Phe Arg Ile Xaa 1 5 <210> SEQ ID NO 73 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 73 Arg Pro Arg Phe Arg Ile Xaa 1 5 <210> SEQ ID NO 74 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 74 Pro Arg Phe Arg Ile Xaa 1 5 <210> SEQ ID NO 75 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-Therapeutic Agent <400> SEQUENCE: 75 Leu Arg Ser Arg Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 76 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 76 Arg Ser Arg Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 77 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 77 Ser Arg Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 78 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is pyroglutamic acid <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 78 Xaa Pro Arg Ala Xaa 1 5 <210> SEQ ID NO 79 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is CH3SO2-D-HHT: HHT is hexahydrotyrosol <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 79 Xaa Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 80 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is N-p-tosyl-Gly <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 80 Xaa Pro Arg Ala Xaa 1 5 <210> SEQ ID NO 81 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Benzoyl-Val <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 81 Xaa Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 82 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is CH3SO2-D-HHT: HHT is hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 82 Xaa Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 83 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is N-aplha-Z-D-Arg: Z is benzyloxycarbonyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 83 Xaa Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 84 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is pyroglutamic acid <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 84 Xaa Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 85 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Ile <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine- therapeutic agent <400> SEQUENCE: 85 Xaa Pro Arg Ala Xaa 1 5 <210> SEQ ID NO 86 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Cbo-L-(gamma)Glu(alpha-t-BuO): Cbo is carbobenzoxy <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 86 Xaa Arg Ala Ala Xaa 1 5 <210> SEQ ID NO 87 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Pro <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 87 Xaa Phe Arg Ala Xaa 1 5 <210> SEQ ID NO 88 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Val <220> FEATURE: <221> NAME/KEY: MUTAGEN <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 88 Xaa Leu Arg Ala Xaa 1 5 <210> SEQ ID NO 89 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Bz-Ile: Bz is benzoyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 89 Xaa Glu Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 90 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Bz-Ile: Bz is benzoyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 90 Xaa Xaa Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 91 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Benzoyl-Pro <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 91 Xaa Phe Arg Ala Xaa 1 5 <210> SEQ ID NO 92 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Phe <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: pipecolinic acid <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 92 Xaa Xaa Arg Ala Xaa 1 5 <210> SEQ ID NO 93 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Val <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 93 Xaa Leu Lys Ala Xaa 1 5 <210> SEQ ID NO 94 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Nle <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: HHT: HHT is hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 94 Xaa Xaa Lys Ala Xaa 1 5 <210> SEQ ID NO 95 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is pyroglutamic acid <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 95 Xaa Arg Thr Lys Arg Ala Xaa 1 5 <210> SEQ ID NO 96 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-Arg <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 96 Xaa Gln Arg Arg Ala Xaa 1 5 <210> SEQ ID NO 97 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Boc-Gln: Boc is t-butoxycarbonyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 97 Xaa Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 98 <211> LENGTH: 4 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Z-Arg: Z is benzyloxycarbonyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 98 Xaa Arg Ala Xaa 1 <210> SEQ ID NO 99 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-HHT: HHT is hexahydrotyrosol <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 99 Xaa Ala Arg Ala Xaa 1 5 <210> SEQ ID NO 100 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-CHT: HHT is hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 100 Xaa Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 101 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is MeSO2-D-Phe <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 101 Xaa Pro Arg Ala Xaa 1 5 <210> SEQ ID NO 102 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is delta-Z-D-Lys: Z is benzyloxycarbonyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 102 Xaa Pro Arg Ala Xaa 1 5 <210> SEQ ID NO 103 <211> LENGTH: 4 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is CH3SO2-D-CHA: CHA is cyclohexylalanyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is But-Arg <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 103 Xaa Xaa Ala Xaa 1 <210> SEQ ID NO 104 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 104 Arg Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 105<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Ala-therapeutic agent <400> SEQUENCE: 105 Arg Arg Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 106<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 106 Leu Arg Arg Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 107 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 107 Arg Gln Ser Arg Xaa 1 5 <210> SEQ ID NO 108<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 108 Arg Arg Gln Ser Arg Xaa 1 5 <210> SEQ ID NO 109<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Glycine-therapeutic agent <400> SEQUENCE: 109 Leu Arg Arg Gln Ser Arg Gly Xaa 1 5 <210> SEQ ID NO 110<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (0)...(0) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 110 Leu Arg Arg Gln Ser Arg Xaa 1 5 <210> SEQ ID NO 111<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (0)...(0) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 111 Arg Arg Gln Ser Arg Xaa 1 5 <210> SEQ ID NO 112<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 112 Leu Arg Arg Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 113 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (1)...(0) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(Alpha-(3-amidinobenzyl) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 113 Leu Arg Ala Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 114 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 114 Leu Arg Ala Xaa Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 115 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 115 Leu Arg Ser Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 116 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 116 Leu Arg Ser Xaa Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 117 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 117 Leu Arg Pro Arg Phe Lys Ser Xaa 1 5 <210> SEQ ID NO 118 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 118 Arg Pro Arg Phe Lys Ser Xaa 1 5 <210> SEQ ID NO 119 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 119 Pro Arg Phe Lys Ser Xaa 1 5 <210> SEQ ID NO 120 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 120 Leu Arg Ser Lys Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 121 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 121 Arg Ser Lys Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 122 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 122 Ser Lys Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 123 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 123 Leu Arg Pro Arg Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 124 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (0)...(0) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 124 Arg Pro Arg Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 125 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 125 Pro Arg Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 126 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (0)...(0) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 126 Leu Arg Ser Arg Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 127 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 127 Arg Ser Arg Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 128 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 128 Ser Arg Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 129 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (0)...(0) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 129 Leu Arg Ala Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 130 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 130 Leu Arg Ala Xaa Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 131 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 131 Leu Arg Ser Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 132 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 132 Leu Arg Ser Xaa Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 133 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 133 Leu Arg Pro Arg Phe Lys Ser Xaa 1 5 <210> SEQ ID NO 134 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 134 Arg Pro Arg Phe Lys Ser Xaa 1 5 <210> SEQ ID NO 135 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 135 Pro Arg Phe Lys Ser Xaa 1 5 <210> SEQ ID NO 136 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 136 Leu Arg Ser Lys Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 137 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 137 Arg Ser Lys Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 138 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 138 Ser Lys Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 139 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 139 Leu Arg Pro Arg Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 140 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 140 Arg Pro Arg Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 141 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 141 Pro Arg Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 142 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 142 Leu Arg Ser Arg Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 143 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 143 Arg Ser Arg Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 144 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 144 Ser Arg Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 145 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is pyroglutamic acid <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 145 Xaa Pro Arg Ser Xaa 1 5 <210> SEQ ID NO 146 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is CH3SO2-D-HHT; HHT is hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 146 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 147 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is n-p-tosyl-Gly <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 147 Xaa Pro Arg Ser Xaa 1 5 <210> SEQ ID NO 148 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Benzoyl-Val <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 148 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 149 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is CH3SO2-D-HHT; HHT is hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 149 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 150 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is N-alpha-Z-D-Arg; Z is benzyloxycarbonyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 150 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 151 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is pyroglutamic acid <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 151 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 152 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Ile <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 152 Xaa Pro Arg Ser Xaa 1 5 <210> SEQ ID NO 153 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Cbo-L-(gamma)Glu(alpha-t-BuO); Cbo is carbobenzoxy <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 153 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 154 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Pro <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 154 Xaa Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 155 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Val <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 155 Xaa Leu Arg Ser Xaa 1 5 <210> SEQ ID NO 156 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Bz-Ile; Bz is benzoyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 156 Xaa Glu Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 157 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Bz-Ile <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Glu(gamma-OMe) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 157 Xaa Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 158 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is benzoyle-Pro <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 158 Xaa Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 159 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Phe <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Pip is pipecolinic acid <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic acid <400> SEQUENCE: 159 Xaa Xaa Arg Ser Xaa 1 5 <210> SEQ ID NO 160 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Val <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic acid <400> SEQUENCE: 160 Xaa Leu Lys Ser Xaa 1 5 <210> SEQ ID NO 161 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Nle <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is HHT: hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Xaa is leucine-therapeutic agent <400> SEQUENCE: 161 Xaa Xaa Lys Ser Xaa 1 5 <210> SEQ ID NO 162 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is pyroglutamic acid <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 162 Xaa Arg Thr Lys Arg Ser Xaa 1 5 <210> SEQ ID NO 163 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-Arg <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 163 Xaa Gln Arg Arg Ser Xaa 1 5 <210> SEQ ID NO 164 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Boc-Gln <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Xaa is Leucine-therapeutic agent <400> SEQUENCE: 164 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 165 <211> LENGTH: 4 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Z-Arg: Z is benzyloxycarbonyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 165 Xaa Arg Ser Xaa 1 <210> SEQ ID NO 166 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-HHT: HHT is hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 166 Xaa Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 167 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-CHT: CHT is hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 167 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 168 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is MeSO2-dPhe <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 168 Xaa Pro Arg Ser Xaa 1 5 <210> SEQ ID NO 169 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is delta-Z-D-Lys: Z is benzyloxycarbonyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 169 Xaa Pro Arg Ser Xaa 1 5 <210> SEQ ID NO 170 <211> LENGTH: 4 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is CH3SO2-D-CHA: CHA is cyclohexylalanyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is But-Arg <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 170 Xaa Xaa Ser Xaa 1 <210> SEQ ID NO 171 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 171 Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 172 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 172 Arg Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 173 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 173 Leu Arg Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 174 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 174 Arg Gln Ser Arg Xaa 1 5 <210> SEQ ID NO 175 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 175 Arg Arg Gln Ser Arg Xaa 1 5 <210> SEQ ID NO 176 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 176 Leu Arg Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 177 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 177 Leu Arg Arg Gln Ser Arg Xaa 1 5 <210> SEQ ID NO 178 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 178 Arg Arg Gln Ser Arg Xaa 1 5 <210> SEQ ID NO 179 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 179 Leu Arg Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 180 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 180 Arg Gln Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 181 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 181 Arg Gln Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 182 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 182 Arg Gln Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 183 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 183 Arg Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 184 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 184 Arg Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 185 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 185 Arg Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 186 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 186 Arg Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 187 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 187 Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 188 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 188 Gln Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 189 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 189 Gln Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 190 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 190 Gln Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 191 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(Alpha-(3-amidinobenzyl) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 191 Leu Arg Ala Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 192 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 192 Leu Arg Ala Xaa Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 193 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 193 Leu Arg Ser Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 194 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 194 Leu Arg Ser Xaa Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 195 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 195 Leu Arg Pro Arg Phe Lys Ser Ser Xaa 1 5 <210> SEQ ID NO 196 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 196 Arg Pro Arg Phe Lys Ser Ser Xaa 1 5 <210> SEQ ID NO 197 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 197 Pro Arg Phe Lys Ser Ser Xaa 1 5 <210> SEQ ID NO 198 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 198 Leu Arg Ser Lys Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 199 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 199 Arg Ser Lys Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 200 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 200 Ser Lys Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 201 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 201 Leu Arg Pro Arg Phe Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 202 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 202 Arg Pro Arg Phe Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 203 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 203 Pro Arg Phe Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 204 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 204 Leu Arg Ser Arg Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 205 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 205 Arg Ser Arg Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 206 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 206 Ser Arg Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 207 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (0)...(0) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 207 Leu Arg Ala Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 208 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 208 Leu Arg Ala Xaa Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 209 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 209 Leu Arg Ser Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 210 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Quat: (R)-Glu(alpha-(3-amidinobenzyl)) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 210 Leu Arg Ser Xaa Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 211 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 211 Leu Arg Pro Arg Phe Lys Ser Ser Xaa 1 5 <210> SEQ ID NO 212 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 212 Arg Pro Arg Phe Lys Ser Ser Xaa 1 5 <210> SEQ ID NO 213 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 213 Pro Arg Phe Lys Ser Ser Xaa 1 5 <210> SEQ ID NO 214 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 214 Leu Arg Ser Lys Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 215 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 215 Arg Ser Lys Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 216 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 216 Ser Lys Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 217 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 217 Leu Arg Pro Arg Phe Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 218 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 218 Arg Pro Arg Phe Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 219 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 219 Pro Arg Phe Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 220 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 220 Leu Arg Ser Arg Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 221 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 221 Arg Ser Arg Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 222 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 222 Ser Arg Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 223 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is pyroglutamic acid <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 223 Xaa Pro Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 224 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is CH3SO2-D-HHT; HHT is hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 224 Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 225 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is n-p-tosyl-Gly <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 225 Xaa Pro Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 226 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Benzoyl-Val <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 226 Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 227 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is CH3SO2-D-HHT; HHT is hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 227 Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 228 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is N-alpha-Z-D-Arg; Z is benzyloxycarbonyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 228 Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 229 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is pyroglutamic acid <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 229 Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 230 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Ile <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 230 Xaa Pro Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 231 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Cbo-L-(gamma)Glu(alpha-t-BuO); Cbo is carbobenzoxy <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 231 Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 232 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Pro <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 232 Xaa Phe Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 233 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Val <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 233 Xaa Leu Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 234 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Bz-Ile; Bz is benzoyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 234 Xaa Glu Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 235 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Bz-Ile <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Glu(gamma-OMe) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 235 Xaa Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 236 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is benzoyle-Pro <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 236 Xaa Phe Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 237 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Phe <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Pip is pipecolinic acid <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic acid <400> SEQUENCE: 237 Xaa Xaa Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 238 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Val <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic acid <400> SEQUENCE: 238 Xaa Leu Lys Ser Ser Xaa 1 5 <210> SEQ ID NO 239 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-Nle <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is HHT: hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Xaa is leucine-therapeutic agent <400> SEQUENCE: 239 Xaa Xaa Lys Ser Ser Xaa 1 5 <210> SEQ ID NO 240 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is pyroglutamic acid <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 240 Xaa Arg Thr Lys Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 241 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-Arg <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 241 Xaa Gln Arg Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 242 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Boc-Gln <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Xaa is Leucine-therapeutic agent <400> SEQUENCE: 242 Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 243 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Z-Arg: Z is benzyloxycarbonyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 243 Xaa Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 244 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-HHT: HHT is hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 244 Xaa Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 245 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is H-D-CHT: CHT is hexahydrotyrosyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 245 Xaa Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 246 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is MeSO2-dPhe <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 246 Xaa Pro Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 247 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is delta-Z-D-Lys: Z is benzyloxycarbonyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 247 Xaa Pro Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 248 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is CH3SO2-D-CHA: CHA is cyclohexylalanyl <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is But-Arg <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 248 Xaa Xaa Ser Ser Xaa 1 5 <210> SEQ ID NO 249 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 249 Arg Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 250 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 250 Arg Arg Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 251 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 251 Leu Arg Arg Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 252 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 252 Arg Gln Ser Arg Xaa 1 5 <210> SEQ ID NO 253 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 253 Arg Arg Gln Ser Arg Xaa 1 5 <210> SEQ ID NO 254 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 254 Leu Arg Arg Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 255 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 255 Leu Arg Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 256 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 256 Arg Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 257 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 257 Leu Arg Arg Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 258 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 258 Arg Gln Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 259 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 259 Arg Gln Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 260 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic acid <400> SEQUENCE: 260 Arg Gln Phe Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 261 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 261 Arg Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 262 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 262 Arg Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 263 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 263 Arg Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 264 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 264 Arg Phe Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 265 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 265 Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 266 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 266 Gln Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 267 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: leucine-therapeutic agent <400> SEQUENCE: 267 Gln Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 268 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 268 Gln Phe Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 269 <211> LENGTH: 816 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (1)...(816) <223> OTHER INFORMATION: Nucleotide sequence encoding MTSP25, including MTSP25 protease domain <220> FEATURE: <221> NAME/KEY: misc_feature <222> LOCATION: (248)...(270) <223> OTHER INFORMATION: Transmembrane domain encompasses amino acids 248-270 at the C-terminus of the trypsin-like serine protease domain (amino acids 1-237) <400> SEQUENCE: 269 att ata ggg ggc acc gaa gca caa gct ggc gca tgg ccg tgg gtg gtg 48 Ile Ile Gly Gly Thr Glu Ala Gln Ala Gly Ala Trp Pro Trp Val Val 1 5 10 15 agc ctg cag att aaa tat ggc cgt gtt ctt gtt cat gta tgt ggg gga 96 Ser Leu Gln Ile Lys Tyr Gly Arg Val Leu Val His Val Cys Gly Gly 20 25 30 acc cta gtg aga gag agg tgg gtc ctc aca gct gcc cac tgc act aaa 144 Thr Leu Val Arg Glu Arg Trp Val Leu Thr Ala Ala His Cys Thr Lys 35 40 45 gac gct agc gat cct tta atg tgg aca gct gtg att gga act aat aat 192 Asp Ala Ser Asp Pro Leu Met Trp Thr Ala Val Ile Gly Thr Asn Asn 50 55 60 ata cat gga cgc tat cct cat acc aag aag ata aaa att aaa gca atc 240 Ile His Gly Arg Tyr Pro His Thr Lys Lys Ile Lys Ile Lys Ala Ile 65 70 75 80 att att cat cca aac ttc att ttg gaa tct tat gta aat gat att gca 288 Ile Ile His Pro Asn Phe Ile Leu Glu Ser Tyr Val Asn Asp Ile Ala 85 90 95 ctt ttt cac tta aaa aaa gca gtg agg tat aat gac tat att cag cct 336Leu Phe His Leu Lys Lys Ala Val Arg Tyr Asn Asp Tyr Ile Gln Pro 100 105 110 att tgc cta cct ttt gat gtt ttc caa atc ctg gac gga aac aca aag 384 Ile Cys Leu Pro Phe Asp Val Phe Gln Ile Leu Asp Gly Asn Thr Lys 115 120 125 tgt ttt ata agt ggc tgg gga aga aca aaa gaa gaa ggt aac gct aca 432 Cys Phe Ile Ser Gly Trp Gly Arg Thr Lys Glu Glu Gly Asn Ala Thr 130 135 140 aat att tta caa gat gca gaa gtg cat tat att tct cga gag atg tgt 480Asn Ile Leu Gln Asp Ala Glu Val His Tyr Ile Ser Arg Glu Met Cys 145 150 155 160 aat tct gag agg agt tat ggg gga ata att cct aac act tca ttt tgt 528Asn Ser Glu Arg Ser Tyr Gly Gly Ile Ile Pro Asn Thr Ser Phe Cys 165 170 175 gca ggt gat gaa gat gga gct ttt gat act tgc agg ggt gac agt ggg 576 Ala Gly Asp Glu Asp Gly Ala Phe Asp Thr Cys Arg Gly Asp Ser Gly 180 185 190 gga cca tta atg tgc tac tta cca gaa tat aaa aga ttt ttt gta atg 624 Gly Pro Leu Met Cys Tyr Leu Pro Glu Tyr Lys Arg Phe Phe Val Met 195 200 205 gga att acc agt tac gga cat ggc tgt ggt cga aga ggt ttt cct ggt 672 Gly Ile Thr Ser Tyr Gly His Gly Cys Gly Arg Arg Gly Phe Pro Gly 210 215 220 gtc tat att ggg cca tcc ttc tac caa aag tgg ctg aca gag cat ttc 720 Val Tyr Ile Gly Pro Ser Phe Tyr Gln Lys Trp Leu Thr Glu His Phe 225 230 235 240 ttc cat gca agc act caa ggc ata ctt act ata aat att tta cgt ggc 768 Phe His Ala Ser Thr Gln Gly Ile Leu Thr Ile Asn Ile Leu Arg Gly 245 250 255 cag atc ctc ata gct tta tgt ttt gtc atc tta cta gca aca aca taa 816 Gln Ile Leu Ile Ala Leu Cys Phe Val Ile Leu Leu Ala Thr Thr * 260 265 270 <210> SEQ ID NO 270 <211> LENGTH: 271 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 270 Ile Ile Gly Gly Thr Glu Ala Gln Ala Gly Ala Trp Pro Trp Val Val 1 5 10 15 Ser Leu Gln Ile Lys Tyr Gly Arg Val Leu Val His Val Cys Gly Gly 20 25 30 Thr Leu Val Arg Glu Arg Trp Val Leu Thr Ala Ala His Cys Thr Lys 35 40 45 Asp Ala Ser Asp Pro Leu Met Trp Thr Ala Val Ile Gly Thr Asn Asn 50 55 60 Ile His Gly Arg Tyr Pro His Thr Lys Lys Ile Lys Ile Lys Ala Ile 65 70 75 80 Ile Ile His Pro Asn Phe Ile Leu Glu Ser Tyr Val Asn Asp Ile Ala 85 90 95 Leu Phe His Leu Lys Lys Ala Val Arg Tyr Asn Asp Tyr Ile Gln Pro 100 105 110 Ile Cys Leu Pro Phe Asp Val Phe Gln Ile Leu Asp Gly Asn Thr Lys 115 120 125 Cys Phe Ile Ser Gly Trp Gly Arg Thr Lys Glu Glu Gly Asn Ala Thr 130 135 140 Asn Ile Leu Gln Asp Ala Glu Val His Tyr Ile Ser Arg Glu Met Cys 145 150 155 160 Asn Ser Glu Arg Ser Tyr Gly Gly Ile Ile Pro Asn Thr Ser Phe Cys 165 170 175 Ala Gly Asp Glu Asp Gly Ala Phe Asp Thr Cys Arg Gly Asp Ser Gly 180 185 190 Gly Pro Leu Met Cys Tyr Leu Pro Glu Tyr Lys Arg Phe Phe Val Met 195 200 205 Gly Ile Thr Ser Tyr Gly His Gly Cys Gly Arg Arg Gly Phe Pro Gly 210 215 220 Val Tyr Ile Gly Pro Ser Phe Tyr Gln Lys Trp Leu Thr Glu His Phe 225 230 235 240 Phe His Ala Ser Thr Gln Gly Ile Leu Thr Ile Asn Ile Leu Arg Gly 245 250 255 Gln Ile Leu Ile Ala Leu Cys Phe Val Ile Leu Leu Ala Thr Thr 260 265 270 <210> SEQ ID NO 271 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: amino acids 401-407 of SEQ ID No. 97 in WO 02/00860 <400> SEQUENCE: 271 Arg Lys His Leu Pro Arg Pro Ala 1 5 <210> SEQ ID NO 272 <211> LENGTH: 228 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: alternative PD1 of MTSP12 <400> SEQUENCE: 272 Ile Val Gly Gly Met Glu Ala Ser Pro Gly Glu Phe Pro Trp Gln Ala 1 5 10 15 Ser Leu Arg Glu Asn Lys Glu His Phe Cys Gly Ala Ala Ile Ile Asn 20 25 30 Ala Arg Trp Leu Val Ser Ala Ala His Cys Phe Asn Glu Phe Gln Asp 35 40 45 Pro Thr Lys Trp Val Ala Tyr Val Gly Ala Thr Tyr Leu Ser Gly Ser 50 55 60 Glu Ala Ser Thr Val Arg Ala Gln Val Val Gln Ile Val Lys His Pro 65 70 75 80 Leu Tyr Asn Ala Asp Thr Ala Asp Phe Asp Val Ala Val Leu Glu Leu 85 90 95 Thr Ser Pro Leu Pro Phe Gly Arg His Ile Gln Pro Val Cys Leu Pro 100 105 110 Ala Ala Thr His Ile Phe Pro Pro Ser Lys Lys Cys Leu Ile Ser Gly 115 120 125 Trp Gly Tyr Leu Lys Glu Asp Phe Leu Arg Lys His Leu Pro Arg Pro 130 135 140 Ala Val Lys Pro Gly Val Leu Gln Lys Ala Thr Val Glu Leu Leu Asp 145 150 155 160 Gln Ala Leu Cys Ala Ser Leu Tyr Gly His Ser Leu Thr Asp Arg Met 165 170 175 Val Cys Ala Gly Tyr Leu Asp Gly Lys Val Asp Ser Cys Gln Gly Asp 180 185 190 Ser Gly Gly Pro Leu Val Cys Glu Glu Pro Ser Gly Arg Phe Ser Leu 195 200 205 Ala Gly Ile Val Ser Trp Gly Ile Gly Cys Ala Glu Ala Arg Arg Pro 210 215 220 Gly Val Tyr Ala 225 <210> SEQ ID NO 273 <211> LENGTH: 804 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (1)...(804) <223> OTHER INFORMATION: Nucleotide sequence encoding MTSP20, including MTSP20 protease domain <400> SEQUENCE: 273 aca gca ggt ccc cag gca gga gca ccc tcc cca tgg ccc tgg gag gcc 48 Thr Ala Gly Pro Gln Ala Gly Ala Pro Ser Pro Trp Pro Trp Glu Ala 1 5 10 15 agg ctg atg cac cag gga cag ctg gcc tgt ggc gga gcc ctg gtg tca 96 Arg Leu Met His Gln Gly Gln Leu Ala Cys Gly Gly Ala Leu Val Ser 20 25 30 gag gag acg gtg cta act gtt gcc cac tgc ttc att ggg cgc cag gcc 144 Glu Glu Thr Val Leu Thr Val Ala His Cys Phe Ile Gly Arg Gln Ala 35 40 45 cca gag gaa tgg agc gta ggg ctg ggg acc aga ccg gag gag tgg ggc 192 Pro Glu Glu Trp Ser Val Gly Leu Gly Thr Arg Pro Glu Glu Trp Gly 50 55 60 ctg aag cag ctc atc ctg cat gga gcc tac acc cac cct gag ggg ggc 240 Leu Lys Gln Leu Ile Leu His Gly Ala Tyr Thr His Pro Glu Gly Gly 65 70 75 80 tac gac atg gcc ctc ctg ctg ctg gcc cag cct gtg aca ctg gga gcc 288Tyr Asp Met Ala Leu Leu Leu Leu Ala Gln Pro Val Thr Leu Gly Ala 85 90 95 agc ctg cgg ccc ctc tgc ctg ccc tat cct gac cac cac ctg cct gat 336 Ser Leu Arg Pro Leu Cys Leu Pro Tyr Pro Asp His His Leu Pro Asp 100 105 110 ggg gag cgt ggc tgg gtt ctg gga cgg gcc cgc cca gga gca ggc atc 384Gly Glu Arg Gly Trp Val Leu Gly Arg Ala Arg Pro Gly Ala Gly Ile 115 120 125 agc tcc ctc cag aca gtg ccc gtg acc ctc ctg ggg cct agg gcc tgc 432Ser Ser Leu Gln Thr Val Pro Val Thr Leu Leu Gly Pro Arg Ala Cys 130 135 140 agc cgg ctg cat gca gct cct ggg ggt gat ggc agc cct att ctg ccg 480Ser Arg Leu His Ala Ala Pro Gly Gly Asp Gly Ser Pro Ile Leu Pro 145 150 155 160 ggg atg gtg tgt acc agt gct gtg ggt gag ctg ccc agc tgt gag ggc 528Gly Met Val Cys Thr Ser Ala Val Gly Glu Leu Pro Ser Cys Glu Gly 165 170 175 ctg tct ggg gca cca ctg gtg cat gag gtg agg ggc aca tgg ttc ctg 576 Leu Ser Gly Ala Pro Leu Val His Glu Val Arg Gly Thr Trp Phe Leu 180 185 190 gcc ggg ctg cac agc ttc gga gat gct tgc caa ggc ccc gcc agg ccg 624 Ala Gly Leu His Ser Phe Gly Asp Ala Cys Gln Gly Pro Ala Arg Pro 195 200 205 gcg gtc ttc acc gcg ctc cct gcc tat gag gac tgg gtc agc agt ttg 672 Ala Val Phe Thr Ala Leu Pro Ala Tyr Glu Asp Trp Val Ser Ser Leu 210 215 220 gac tgg cag gtc tac ttc gcc gag gaa cca gag ccc gag gct gag cct 720 Asp Trp Gln Val Tyr Phe Ala Glu Glu Pro Glu Pro Glu Ala Glu Pro 225 230 235 240 gga agc tgc ctg gcc aac atg agt atg tgg ccc cgg ggc ctc ctg cca 768 Gly Ser Cys Leu Ala Asn Met Ser Met Trp Pro Arg Gly Leu Leu Pro 245 250 255 aac cct gcc tct cca gga ccc ttc tct ctc cag tga 804 Asn Pro Ala Ser Pro Gly Pro Phe Ser Leu Gln * 260 265 <210> SEQ ID NO 274 <211> LENGTH: 267 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 274 Thr Ala Gly Pro Gln Ala Gly Ala Pro Ser Pro Trp Pro Trp Glu Ala 1 5 10 15 Arg Leu Met His Gln Gly Gln Leu Ala Cys Gly Gly Ala Leu Val Ser 20 25 30 Glu Glu Thr Val Leu Thr Val Ala His Cys Phe Ile Gly Arg Gln Ala 35 40 45 Pro Glu Glu Trp Ser Val Gly Leu Gly Thr Arg Pro Glu Glu Trp Gly 50 55 60 Leu Lys Gln Leu Ile Leu His Gly Ala Tyr Thr His Pro Glu Gly Gly 65 70 75 80 Tyr Asp Met Ala Leu Leu Leu Leu Ala Gln Pro Val Thr Leu Gly Ala 85 90 95 Ser Leu Arg Pro Leu Cys Leu Pro Tyr Pro Asp His His Leu Pro Asp 100 105 110 Gly Glu Arg Gly Trp Val Leu Gly Arg Ala Arg Pro Gly Ala Gly Ile 115 120 125 Ser Ser Leu Gln Thr Val Pro Val Thr Leu Leu Gly Pro Arg Ala Cys 130 135 140 Ser Arg Leu His Ala Ala Pro Gly Gly Asp Gly Ser Pro Ile Leu Pro 145 150 155 160 Gly Met Val Cys Thr Ser Ala Val Gly Glu Leu Pro Ser Cys Glu Gly 165 170 175 Leu Ser Gly Ala Pro Leu Val His Glu Val Arg Gly Thr Trp Phe Leu 180 185 190 Ala Gly Leu His Ser Phe Gly Asp Ala Cys Gln Gly Pro Ala Arg Pro 195 200 205 Ala Val Phe Thr Ala Leu Pro Ala Tyr Glu Asp Trp Val Ser Ser Leu 210 215 220 Asp Trp Gln Val Tyr Phe Ala Glu Glu Pro Glu Pro Glu Ala Glu Pro 225 230 235 240 Gly Ser Cys Leu Ala Asn Met Ser Met Trp Pro Arg Gly Leu Leu Pro 245 250 255 Asn Pro Ala Ser Pro Gly Pro Phe Ser Leu Gln 260 265 <210> SEQ ID NO 275 <211> LENGTH: 699 <212> TYPE: DNA <213> ORGANISM: Homo Sapien <220> FEATURE: <221> NAME/KEY: CDS <222> LOCATION: (1)...(699) <223> OTHER INFORMATION: Nucleotide sequence encoding MTSP22, including MTSP22 protease domain <400> SEQUENCE: 275 att gtg aat gga aaa agc tcc ctg gag ggg gca tgg cca tgg cag gcc 48 Ile Val Asn Gly Lys Ser Ser Leu Glu Gly Ala Trp Pro Trp Gln Ala 1 5 10 15 agc atg caa tgg aaa ggc cgt cac tac tgt gga gcc tct ctg atc agc 96 Ser Met Gln Trp Lys Gly Arg His Tyr Cys Gly Ala Ser Leu Ile Ser 20 25 30 agc agg tgg cta tta tct gca gct cac tgc ttt gct aag aaa aat aat 144 Ser Arg Trp Leu Leu Ser Ala Ala His Cys Phe Ala Lys Lys Asn Asn 35 40 45 tca aaa gat tgg act gtc aac ttt gga gtt gta gta aat aaa cca tat 192 Ser Lys Asp Trp Thr Val Asn Phe Gly Val Val Val Asn Lys Pro Tyr 50 55 60 atg aca cgg aaa gtc caa aac att att ttt cat gaa aat tat agc agt 240 Met Thr Arg Lys Val Gln Asn Ile Ile Phe His Glu Asn Tyr Ser Ser 65 70 75 80 cct ggg ctt cat gat gat att gcc ctt gtg cag ctt gct gaa gaa gtt 288Pro Gly Leu His Asp Asp Ile Ala Leu Val Gln Leu Ala Glu Glu Val 85 90 95 tct ttt aca gag tac att cgt aag att tgt ctt cct gaa gcc aaa atg 336Ser Phe Thr Glu Tyr Ile Arg Lys Ile Cys Leu Pro Glu Ala Lys Met 100 105 110 aag ctc tca gaa aat gac aat gtt gta gtt aca ggt tgg gga aca ctt 384Lys Leu Ser Glu Asn Asp Asn Val Val Val Thr Gly Trp Gly Thr Leu 115 120 125 tat atg aat ggt tca ttt cca gtg ata ctt caa gaa gcc ttt ttg aag 432Tyr Met Asn Gly Ser Phe Pro Val Ile Leu Gln Glu Ala Phe Leu Lys 130 135 140 att att gac aac aaa att tgc aat gcc tca tat gca tac tct ggc tta 480Ile Ile Asp Asn Lys Ile Cys Asn Ala Ser Tyr Ala Tyr Ser Gly Leu 145 150 155 160 gtg act gat aca atg tta tgt gct gga ttt atg tca gga gaa gct gat 528 Val Thr Asp Thr Met Leu Cys Ala Gly Phe Met Ser Gly Glu Ala Asp 165 170 175 gca tgt cag aat gat tct ggt gga cca cta gct tac cct gat tcc aga 576 Ala Cys Gln Asn Asp Ser Gly Gly Pro Leu Ala Tyr Pro Asp Ser Arg 180 185 190 aat atc tgg cat ctt gtt gga ata gta agc tgg ggt gat gga tgt ggt 624 Asn Ile Trp His Leu Val Gly Ile Val Ser Trp Gly Asp Gly Cys Gly 195 200 205 aaa aag aat aag cca ggt gtc tat act cga gtg act tct tat cgc aat 672 Lys Lys Asn Lys Pro Gly Val Tyr Thr Arg Val Thr Ser Tyr Arg Asn 210 215 220 tgg att aca tcc aag act gga ctc tga 699 Trp Ile Thr Ser Lys Thr Gly Leu * 225 230 <210> SEQ ID NO 276 <211> LENGTH: 232 <212> TYPE: PRT <213> ORGANISM: Homo Sapien <400> SEQUENCE: 276 Ile Val Asn Gly Lys Ser Ser Leu Glu Gly Ala Trp Pro Trp Gln Ala 1 5 10 15 Ser Met Gln Trp Lys Gly Arg His Tyr Cys Gly Ala Ser Leu Ile Ser 20 25 30 Ser Arg Trp Leu Leu Ser Ala Ala His Cys Phe Ala Lys Lys Asn Asn 35 40 45 Ser Lys Asp Trp Thr Val Asn Phe Gly Val Val Val Asn Lys Pro Tyr 50 55 60 Met Thr Arg Lys Val Gln Asn Ile Ile Phe His Glu Asn Tyr Ser Ser 65 70 75 80 Pro Gly Leu His Asp Asp Ile Ala Leu Val Gln Leu Ala Glu Glu Val 85 90 95 Ser Phe Thr Glu Tyr Ile Arg Lys Ile Cys Leu Pro Glu Ala Lys Met 100 105 110 Lys Leu Ser Glu Asn Asp Asn Val Val Val Thr Gly Trp Gly Thr Leu 115 120 125 Tyr Met Asn Gly Ser Phe Pro Val Ile Leu Gln Glu Ala Phe Leu Lys 130 135 140 Ile Ile Asp Asn Lys Ile Cys Asn Ala Ser Tyr Ala Tyr Ser Gly Leu 145 150 155 160 Val Thr Asp Thr Met Leu Cys Ala Gly Phe Met Ser Gly Glu Ala Asp 165 170 175 Ala Cys Gln Asn Asp Ser Gly Gly Pro Leu Ala Tyr Pro Asp Ser Arg 180 185 190 Asn Ile Trp His Leu Val Gly Ile Val Ser Trp Gly Asp Gly Cys Gly 195 200 205 Lys Lys Asn Lys Pro Gly Val Tyr Thr Arg Val Thr Ser Tyr Arg Asn 210 215 220 Trp Ile Thr Ser Lys Thr Gly Leu <210> SEQ ID NO 277<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 277 Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 278<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Xaa is Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 278 Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 279<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 279 Gly Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 280<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 280 Gly Ser Gly Arg Xaa 1 5 <210> SEQ ID NO 281<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: AMIDATION <222> LOCATION: 6 <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is 4-Guanidino-phenylglycine <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 281 Gly Ser Gly Xaa Ser Leu 1 5 <210> SEQ ID NO 282<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Cyclohexylamine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 282 Gly Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 283<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 283 Gly Ser Gly Arg Ala Ser Xaa 1 5 <210> SEQ ID NO 284<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 284 Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 285<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (0)...(0) <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 285 Gly Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 286<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Succinyl-BAlanine <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 286 Ala Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 287<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 287 Gly Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 288<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Homoserine <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-Therapeutic AgentNle <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 288 Gly Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 289<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is D Serine <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 289 Gly Xaa Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 290<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 290 Gly Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 291<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 291 Gly Ser Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 292<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 292 Gly Ser Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 293<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 293 Gly Ser Ala Arg Ala Ser Xaa 1 5 <210> SEQ ID NO 294<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 294 Val Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 295<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 295 Val Ser Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 296<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 296 Val Ser Gly Arg Ala Ser Xaa 1 5 <210> SEQ ID NO 297<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 297 Val Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 298<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 298 Val Ser Ala Arg Met Xaa 1 5 <210> SEQ ID NO 299<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Xaa is Nle <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 299 Val Ser Ala Arg Xaa Xaa 1 5 <210> SEQ ID NO 300<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 300 Val Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 301<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 301 Val Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 302<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is S-Methylcysteine <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: dValine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 302 Xaa Pro Gly Arg Val Xaa 1 5 <210> SEQ ID NO 303<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is S-Methylcysteine <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Valine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 303 Xaa Pro Gly Arg Val Xaa 1 5 <210> SEQ ID NO 304<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is S-Methylcysteine <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 304 Xaa Pro Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 305<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is S-Methylcysteine <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 305 Xaa Pro Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 306<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is S-Methylcysteine <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 306 Xaa Pro Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 307<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is S-Methylcysteine <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 307 Xaa Pro Ala Arg Ala Ser Xaa 1 5 <210> SEQ ID NO 308<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is t-Butyl Glycine <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 308 Xaa Pro Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 309<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 3 <223> OTHER INFORMATION: Xaa is D Serine <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 309 Arg Gly Xaa Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 310<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 310 Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 311<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 311 Arg Gly Ser Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 312<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 312 Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 313<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 313 Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 314<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 314 Arg Gly Ser Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 315<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 315 Arg Gly Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 316<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 316 Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 317<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 317 Arg Gly Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 318<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 318 Arg Gly Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 319<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 319 Arg Gly Ser Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 320<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Nva-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 320 Arg Gly Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 321<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Nva-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 321 Arg Gly Ser Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 322<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 322 Arg Gly Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 323<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is S-MethylCysteine <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Valine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 323 Arg Xaa Pro Gly Arg Val Xaa 1 5 <210> SEQ ID NO 324<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is S-Methylcysteine <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Valine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 324 Arg Xaa Pro Gly Arg Val Xaa 1 5 <210> SEQ ID NO 325<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is S-Methylcysteine <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 325 Arg Xaa Pro Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 326<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 326 Arg Leu Pro Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 327<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: VARIANT <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 327 Arg Val Pro Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 328<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: VARIANT <222> LOCATION: 8 <223> OTHER INFORMATION: dLeucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 328 Arg Val Pro Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 329<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Nle <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 329 Arg Xaa Pro Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 330<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is t-Butylglycine <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 330 Arg Xaa Pro Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 331<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 331 Arg Leu Pro Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 332<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 332 Arg Val Pro Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 333<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Nle <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 333 Arg Xaa Pro Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 334<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 334 Ile Val Ser Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 335<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 335 Ile Val Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 336<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 336 Ile Val Ser Gly Arg Ala Ser Xaa 1 5 <210> SEQ ID NO 337<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 337 Ile Val Ser Ala Arg Met Xaa 1 5 <210> SEQ ID NO 338<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Xaa is Nle <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 338 Ile Val Ser Ala Arg Xaa Xaa 1 5 <210> SEQ ID NO 339<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 339 Ile Val Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 340<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Nle-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 340 Ile Val Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 341<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 341 Ile Val Ser Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 342<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: VARIANT <222> LOCATION: 8 <223> OTHER INFORMATION: dLeucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 342 Leu Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 343<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 343 Leu Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 344<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 344 Leu Arg Gly Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 345<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 345 Leu Arg Gly Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 346<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 346 Leu Arg Gly Ser Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 347<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 347 Leu Arg Gly Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 348<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 348 Leu Arg Gly Ser Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 349<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Nva-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 349 Leu Arg Gly Ser Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 350<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 350 Leu Arg Gly Ser Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 351<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 351 Val Ile Val Ser Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 352<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 352 Val Ile Val Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 353<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 353 Val Ile Val Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 354<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 354 Val Ile Val Ser Ala Arg Met Xaa 1 5 <210> SEQ ID NO 355<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Xaa is Nle <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 355 Val Ile Val Ser Ala Arg Xaa Xaa 1 5 <210> SEQ ID NO 356<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 356 Val Ile Val Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 357<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: dCyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 357 Val Ile Val Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 358<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 358 Val Ile Val Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 359<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 359 Val Ile Val Ser Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 360<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 3 <223> OTHER INFORMATION: Xaa is S-methylcysteine <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Valine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 360 Arg Arg Xaa Pro Gly Arg Val Xaa 1 5 <210> SEQ ID NO 361<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 3 <223> OTHER INFORMATION: Xaa is Nva <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 361 Arg Arg Xaa Pro Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 362<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 362 Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 363<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 363 Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 364<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 364 Ser Gly Arg Ser Ser Ser Xaa 1 5 <210> SEQ ID NO 365<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 365 Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 366<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: dNva-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 366 Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 367<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nva-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 367 Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 368<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (0)...(0) <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Hexylglycine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 368 Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 369<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 369 Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 370<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Homocyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 370 Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 371<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 371 Ser Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 372<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 372 Ser Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 373<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 373 Ser Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 374<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Abu-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 374 Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 375<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 375 Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 376<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nva-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 376 Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 377<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 377 Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 378<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Hexylglycine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 378 Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 379<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 379 Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 380<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Homocyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 380 Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 381<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Abu-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 381 Thr Gly Arg Thr Xaa 1 5 <210> SEQ ID NO 382<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Homoserine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 382 Thr Gly Arg Xaa Xaa 1 5 <210> SEQ ID NO 383<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Abu <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 383 Thr Gly Arg Xaa Xaa 1 5 <210> SEQ ID NO 384<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Abu <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nva-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 384 Thr Gly Arg Xaa Xaa 1 5 <210> SEQ ID NO 385<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 3 <223> OTHER INFORMATION: Xaa is 4-Guanidinophenylalanine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 385 Thr Gly Xaa Ser Xaa 1 5 <210> SEQ ID NO 386<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 3 <223> OTHER INFORMATION: Xaa is 4-Guanidinophenylalanine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 386 Thr Gly Xaa Ser Xaa 1 5 <210> SEQ ID NO 387<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 3 <223> OTHER INFORMATION: Xaa is 4-Guanidinophenylalanine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Abu <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nva-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 387 Thr Gly Xaa Xaa Xaa 1 5 <210> SEQ ID NO 388<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 3 <223> OTHER INFORMATION: Xaa is Alloc <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 388 Thr Gly Xaa Ser Xaa 1 5 <210> SEQ ID NO 389<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 389 Thr Gly Lys Ser Xaa 1 5 <210> SEQ ID NO 390<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 3 <223> OTHER INFORMATION: Xaa is Homoarginine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 390 Thr Gly Xaa Ser Xaa 1 5 <210> SEQ ID NO 391<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is N-homoserine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 391 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 392<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is N-Methyloxycarbonyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 392 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 393<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Phenylsulfonyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 393 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 394<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxyethylcarbonyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 394 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 395<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxydiethoxyacetyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 395 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 396<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is 4-Oxo-pentanoyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 396 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 397 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is 2-Benzo[1,3]dioxo1-5-yl acetylthreonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 397 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 398<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is 2-Pyridin-2-yl-acetyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 398 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 399<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Benzoylacetyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 399 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 400<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is 2-Hydroxy-3-phenyl propionylacetyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate SEQUENCE: 400 Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 401<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxyacetylthreonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 401 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 402<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Phenylacetyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 402 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 403<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is 3-Methoxypropionyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 403 Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 404<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxyethoxyacetyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 404 Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 405<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is 1-Carboxybutanoyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 405 Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 406<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Carboxybenzyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 406 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 407<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Ethoxycarbonylthreonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 407 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 408<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is BAlanine <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 408 Xaa Thr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 409<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Pent-4-ynoyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 409 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 410<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Naphthaacetyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 410 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 411<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Isobutyloxycarbonyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjuagate <400> SEQUENCE: 411 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 412<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Hydroxyacetyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 412 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 413<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarboxylpropanoyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 413 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 414<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is N,N-dimethyl glycine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 414 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 415<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Succinyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 415 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 416<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Formyl threonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 416 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 417<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 417 Thr Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 418<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 3 <223> OTHER INFORMATION: Xaa is 4-Guanidinophenylalanine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 418 Thr Ala Xaa Ser Xaa 1 5 <210> SEQ ID NO 419<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is Abu <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nva-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 419 Thr Ala Arg Xaa Xaa 1 5 <210> SEQ ID NO 420<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Abu-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 420 Thr Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 421<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Abu-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 421 Thr Ala Arg Thr Xaa 1 5 <210> SEQ ID NO 422<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Serine methyl ester <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 422 Thr Xaa Arg Ser Xaa 1 5 <210> SEQ ID NO 423<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Homoserine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 423 Thr Xaa Arg Ser Xaa 1 5 <210> SEQ ID NO 424<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is 1-Methyl histidine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 424 Thr Xaa Arg Ser Xaa 1 5 <210> SEQ ID NO 425<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is 3-Methyl histidine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 425 Thr Xaa Arg Ser Xaa 1 5 <210> SEQ ID NO 426<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 426 Thr His Arg Ser Xaa 1 5 <210> SEQ ID NO 427<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is MeGlycine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 427 Thr Xaa Arg Ser Xaa 1 5 <210> SEQ ID NO 428<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Nva <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 428 Thr Xaa Arg Ser Xaa 1 5 <210> SEQ ID NO 429<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Nle <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 429 Thr Xaa Arg Ser Xaa 1 5 <210> SEQ ID NO 430<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Cyclohexyl alanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 430 Thr Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 431<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Abu <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 431 Thr Xaa Arg Ser Xaa 1 5 <210> SEQ ID NO 432<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is 4,4-Dimethylthreonine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 432 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 433<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Homoserine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 433 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 434<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1,4 <223> OTHER INFORMATION: Xaa is Homoserine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Cyclohexyl alanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 434 Xaa Gly Arg Xaa Xaa 1 5 <210> SEQ ID NO 435<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Homoserine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 435 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 436<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Homoserine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 436 Xaa Gly Arg Thr Xaa 1 5 <210> SEQ ID NO 437<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Homoserine <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 437 Xaa Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 438<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 438 Asn Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 439<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 439 Tyr Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 440<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 440 Tyr Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 441<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 441 Gln Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 442<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nva-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 442 Gln Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 443 <211> LENGTH: 4 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is N-homoserine <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Nle-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 443 Xaa Arg Ser Xaa 1 <210> SEQ ID NO 444 <211> LENGTH: 4 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is N-homoserine <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Nva-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 444 Xaa Arg Ser Xaa 1 <210> SEQ ID NO 445<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: BLOCKED <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 445 Gln Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 446<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 446 Gln Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 447<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 447 Gln Gly Arg Ala Ser Xaa 1 5 <210> SEQ ID NO 448<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic Agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 448 Asn Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 449<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nleucine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 449 Gln Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 450<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nvaline-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 450 Gln Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 451<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <220> FEATURE: <223> OTHER INFORMATION: conjugate <400> SEQUENCE: 451 Gln Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 452<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Allyl-therapeutic agent <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: conjugate <220> FEATURE: <223> OTHER INFORMATION: Xaa is dSerine <400> SEQUENCE: 452 Gln Gly Arg Ser Xaa Xaa 1 5 <210> SEQ ID NO 453<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Allyl-therapeutic agent <400> SEQUENCE: 453 Gln Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 454<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (0)...(0) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 454 Gln Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 455<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 455 Gln Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 456<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 456 Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 457<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nva-therapeutic agent <400> SEQUENCE: 457 Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 458<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 458 Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 459<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 459 Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 460<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 460 Gln Thr Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 461<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Aib <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 461 Gln Xaa Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 462<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Aib <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 462 Gln Xaa Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 463<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (0)...(0) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Abu <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 463 Gln Xaa Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 464<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Abu <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 464 Gln Xaa Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 465<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Cyclohexylalanine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 465 Gln Xaa Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 466<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 466 Gln Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 467<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 467 Gln Phe Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 468<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 468 Gln Tyr Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 469<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 469 Arg Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 470<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 470 Arg Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 471<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 471 Arg Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 472<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 472 Arg Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 473<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 473 Arg Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 474<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 474 Arg Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 475<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 475 Arg Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 476<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 476 Arg Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 477<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 477 Arg Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 478<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 478 Arg Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 479<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 479 Arg Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 480<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 480 Arg Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 481<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 481 Tyr Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 482<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is S-Dioxomethionine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 482 Xaa Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 483<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl-(alpha-(3-cyanobenzyl) ) glutamic acid-delta-methyl ester <220> FEATURE: <221> NAME/KEY: AMIDATION <222> LOCATION: 5 <400> SEQUENCE: 483 Xaa Gly Arg Ser Leu 1 5 <210> SEQ ID NO 484<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl-(alpha- (3-amidinobenzyl) ) glutamic acid -delta-methyl ester <220> FEATURE: <221> NAME/KEY: AMIDATION <222> LOCATION: 5 <400> SEQUENCE: 484 Xaa Gly Arg Ser Leu 1 5 <210> SEQ ID NO 485<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl- (alpha-(3-amidinobenzyl)) glutamic acid <220> FEATURE: <221> NAME/KEY: AMIDATION <222> LOCATION: 5 <400> SEQUENCE: 485 Xaa Gly Arg Ser Leu 1 5 <210> SEQ ID NO 486<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl-(alpha-(3-Methylbenzyl) )glutamic acid -delta-methyl ester <220> FEATURE: <221> NAME/KEY: AMIDATION <222> LOCATION: 5 <400> SEQUENCE: 486 Xaa Gly Arg Ser Leu 1 5 <210> SEQ ID NO 487<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl- (alpha-(3-methylbenzyl)) glutamic acid <220> FEATURE: <221> NAME/KEY: AMIDATION <222> LOCATION: 5 <400> SEQUENCE: 487 Xaa Gly Arg Ser Leu 1 5 <210> SEQ ID NO 488<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl-(alpha-(3-cyanobenzyl) ) glutamic acid-delta-methyl ester <220> FEATURE: <221> NAME/KEY: AMIDATION <222> LOCATION: 6 <400> SEQUENCE: 488 Xaa Gly Arg Ser Ser Leu 1 5 <210> SEQ ID NO 489<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl-(alpha-(3-methylbenzyl) )glutamic acid -delta-methyl ester <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 489 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 490<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl-(alpha-(3-cyanobenzyl) ) glutamic acid -delta-methyl ester <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 490 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 491<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 491 Arg Gln Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 492<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 492 Arg Gln Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 493<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 493 Arg Gln Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 494<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nva-therapeutic agent <400> SEQUENCE: 494 Arg Gln Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 495<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Phenylalanine-therapeutic agent <400> SEQUENCE: 495 Arg Gln Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 496<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: dLeucine-therapeutic agent <400> SEQUENCE: 496 Arg Gln Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 497<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 497 Arg Gln Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 498<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: dNle-therapeutic agent <400> SEQUENCE: 498 Arg Gln Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 499<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 499 Arg Gln Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 500<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nva-therapeutic agent <400> SEQUENCE: 500 Arg Gln Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 501<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 501 Arg Gln Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 502<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Phenylalanine-therapeutic agent <400> SEQUENCE: 502 Arg Gln Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 503<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: leucine-therapeutic agent <400> SEQUENCE: 503 Arg Asn Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 504<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 504 Arg Asn Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 505<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 505 Arg Gln Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 506<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 506 Arg Gln Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 507<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nva-therapeutic agent <400> SEQUENCE: 507 Arg Gln Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 508<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 508 Arg Gln Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 509<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 509 Arg Gln Ala Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 510<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 510 Arg Gln Ala Arg Thr Xaa 1 5 <210> SEQ ID NO 511<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 511 Arg Gln Ala Arg Ala Xaa 1 5 <210> SEQ ID NO 512<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 512 Arg Gln Ala Arg Ala Xaa 1 5 <210> SEQ ID NO 513<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nva-therapeutic agent <400> SEQUENCE: 513 Arg Gln Ala Arg Ala Xaa 1 5 <210> SEQ ID NO 514<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 514 Arg Gln Ala Arg Ala Xaa 1 5 <210> SEQ ID NO 515<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 515 Arg Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 516<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 516 Arg Gln Ser Arg Xaa 1 5 <210> SEQ ID NO 517<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 517 Arg Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 518<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 518 Arg Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 519<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 519 Arg Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 520<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nva-therapeutic agent <400> SEQUENCE: 520 Arg Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 521<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 521 Arg Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 522<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 522 Arg Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 523<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 523 Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 524<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: dNle-therapeutic agent <400> SEQUENCE: 524 Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 525<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 525 Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 526<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nva-therapeutic agent <400> SEQUENCE: 526 Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 527<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Allylglycine-therapeutic agent <400> SEQUENCE: 527 Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 528<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 528 Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 529<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle <400> SEQUENCE: 529 Arg Gln Ser Arg Thr Xaa 1 5 <210> SEQ ID NO 530<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 530 Arg Gln Thr Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 531<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 531 Arg Gln Thr Arg Ser Xaa 1 5 <210> SEQ ID NO 532<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 532 Arg Asn Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 533<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 533 Arg Gln Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 534<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: dNle-therapeutic agent <400> SEQUENCE: 534 Arg Gln Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 535<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nva-therapeutic agent <400> SEQUENCE: 535 Arg Gln Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 536<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 536 Arg Gln Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 537<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 537 Arg Gln Phe Arg Ser Xaa 1 5 <210> SEQ ID NO 538<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 538 Arg Gln Phe Arg Ala Xaa 1 5 <210> SEQ ID NO 539<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 539 Arg Gln Phe Arg Ala Xaa 1 5 <210> SEQ ID NO 540<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nva-therapeutic agent <400> SEQUENCE: 540 Arg Gln Phe Arg Ala Xaa 1 5 <210> SEQ ID NO 541<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Cyclohexylalanine-therapeutic agent <400> SEQUENCE: 541 Arg Gln Phe Arg Ala Xaa 1 5 <210> SEQ ID NO 542<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 542 Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 543<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 543 Arg Arg Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 544<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 544 Arg Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 545<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 545 Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 546<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 546 Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 547<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Nle-therapeutic agent <400> SEQUENCE: 547 Arg Gly Ser Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 548<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 548 Arg Gly Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 549<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (0)...(0) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 549 Ile Val Ser Gly Arg Ala Ser Xaa 1 5 <210> SEQ ID NO 550<211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 550 Leu Arg Arg Gln Ser Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 551<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 551 Leu Arg Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 552<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 552 Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 553<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 553 Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 554<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Glycine-therapeutic agent <400> SEQUENCE: 554 Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 555<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 555 Arg Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 556<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 556 Arg Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 557<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alalnine-therapeutic agent <400> SEQUENCE: 557 Arg Gln Ser Arg Ser Xaa 1 5 <210> SEQ ID NO 558<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 558 Arg Gln Ser Arg Ser Ala Xaa 1 5 <210> SEQ ID NO 559<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 559 Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 560<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 560 Ser Gly Arg Ala Xaa 1 5 <210> SEQ ID NO 561<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 561 Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 562<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 562 Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 563<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 563 Ser Gly Arg Ala Ser Xaa 1 5 <210> SEQ ID NO 564<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Glycine-therapeutic agent <400> SEQUENCE: 564 Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 565<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Glycine-therapeutic agent <400> SEQUENCE: 565 Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 566<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 566 Ser Gly Arg Ser Gly Xaa 1 5 <210> SEQ ID NO 567<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Glycine-therapeutic agent <400> SEQUENCE: 567 Ser Gly Arg Ser Gly Xaa 1 5 <210> SEQ ID NO 568<211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Glycine-therapeutic agent <400> SEQUENCE: 568 Gly Thr Gly Arg Ser Gly Xaa 1 5 <210> SEQ ID NO 569 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is D- Serine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 569 Gly Xaa Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 570 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 3 <223> OTHER INFORMATION: Xaa is D-Serine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 570 Arg Gly Xaa Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 571 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 571 Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 572 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 572 Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 573<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 573 Leu Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 574<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 4 <223> OTHER INFORMATION: Xaa is D-Serine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 574 Leu Arg Gly Xaa Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 575 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is S-Methylcysteine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Valine-therapeutic agent <400> SEQUENCE: 575 Xaa Pro Gly Arg Val Xaa 1 5 <210> SEQ ID NO 576 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is S-Methylcysteine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Valine-therapeutic agent <400> SEQUENCE: 576 Xaa Pro Gly Arg Val Xaa 1 5 <210> SEQ ID NO 577 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is S-Methylcysteine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Valine-therapeutic agent <400> SEQUENCE: 577 Arg Xaa Pro Gly Arg Val Xaa 1 5 <210> SEQ ID NO 578 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 3 <223> OTHER INFORMATION: Xaa is S-Methylcysteine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Valine-therapeutic agent <400> SEQUENCE: 578 Arg Arg Xaa Pro Gly Arg Val Xaa 1 5 <210> SEQ ID NO 579 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 579 Val Ser Ala Arg Met Xaa 1 5 <210> SEQ ID NO 580 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 580 Ile Val Ser Ala Arg Met Xaa 1 5 <210> SEQ ID NO 581 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 581 Val Ile Val Ser Ala Arg Met Xaa 1 5 <210> SEQ ID NO 582 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Xaa is Nle <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 582 Val Ile Val Ser Ala Arg Xaa Xaa 1 5 <210> SEQ ID NO 583 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Xaa is Nle <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 583 Val Ser Ala Arg Xaa Xaa 1 5 <210> SEQ ID NO 584 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Xaa is Nle <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 584 Ile Val Ser Ala Arg Xaa Xaa 1 5 <210> SEQ ID NO 585<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 585 Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 586 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 586 Gly Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 587 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 587 Gly Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 588 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 588 Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 589 <211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 589 Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 590 <211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 590 Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 591 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 591 Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 592 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 592 Arg Gly Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 593 <211> LENGTH: 7 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 7 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 593 Arg Gly Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 594 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 594 Leu Arg Gly Ser Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 595 <211> LENGTH: 9 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 9 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 595 Leu Arg Gly Ser Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 596 <211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 596 Leu Arg Gly Ser Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 597<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Alanine-therapeutic agent <400> SEQUENCE: 597 Leu Arg Arg Gln Ser Arg Ala Xaa 1 5 <210> SEQ ID NO 598<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is N-Methylsulfonyl-alpha-cyclohexyl-D- Alanine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Abu <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 598 Xaa Xaa Arg Ser Xaa 1 5 <210> SEQ ID NO 599<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 599 Arg Ala Arg Ser Xaa 1 5 <210> SEQ ID NO 600<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa si Alpha-Cyclohexyl-D-alanine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Abu <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 600 Xaa Xaa Arg Ser Xaa 1 5 <210> SEQ ID NO 601<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Alpha-Cyclohexyl-D-Alanine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Abu <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 601 Xaa Xaa Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 602<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 602 Gln Gly Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 603<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl-D-homophenylalanine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is 4Hyp <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 603 Xaa Xaa Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 604<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl-(alpha)-3-methylbenzyl glutamic acid -delta-methyl ester <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 604 Xaa Gly Arg Ser Xaa 1 5 <210> SEQ ID NO 605<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is D-cyclohexylalanine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is 4Hyp <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 5 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 605 Xaa Xaa Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 606<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is D-Clohexylalanine <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 2 <223> OTHER INFORMATION: Xaa is Abu <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Leucine-therapeutic agent <400> SEQUENCE: 606 Xaa Xaa Arg Ser Ser Xaa 1 5 <210> SEQ ID NO 607<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl-(alpha-(3-cyanobenzyl)) glutamic acid -delta-methyl ester <220> FEATURE: <221> NAME/KEY: AMIDATION <222> LOCATION: 5 <400> SEQUENCE: 607 Xaa Gly Arg Ser Leu 1 5 <210> SEQ ID NO 608<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl-(alpha- (3-amidinobenzyl)) glutamic acid -delta-methyl ester <220> FEATURE: <221> NAME/KEY: AMIDATION <222> LOCATION: 5 <400> SEQUENCE: 608 Xaa Gly Arg Ser Leu 1 5 <210> SEQ ID NO 609<211> LENGTH: 5 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: conjugate <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 1 <223> OTHER INFORMATION: Xaa is Methoxycarbonyl-(alpha- (3-amidinobenzyl)) glutamic acid <220> FEATURE: <221> NAME/KEY: AMIDATION <222> LOCATION: 5 <400> SEQUENCE: 609 Xaa Gly Arg Ser Leu 1 5 <210> SEQ ID NO 610<211> LENGTH: 6 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: (0)...(0) <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 6 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 610 Arg Arg Gln Ser Arg Xaa 1 5 <210> SEQ ID NO 611<211> LENGTH: 8 <212> TYPE: PRT <213> ORGANISM: Artificial Sequence <220> FEATURE: <223> OTHER INFORMATION: Conjugate <220> FEATURE: <221> NAME/KEY: ACETYLATION <222> LOCATION: 1 <220> FEATURE: <221> NAME/KEY: MOD_RES <222> LOCATION: 8 <223> OTHER INFORMATION: Isoleucine-therapeutic agent <400> SEQUENCE: 611 Leu Arg Arg Gln Ser Arg Ala Xaa 1 5
Claims (136)
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